CN107995911B - Benzoxazepine compounds and methods of use thereof - Google Patents

Abstract

Benzoxazepines having phosphoinositide-3-kinase (PI3K) modulating activity or function are described

Oxazolidinone compounds, and stereoisomers, tautomers, or pharmaceutically acceptable salts thereof, selected from the structures of formula I, below, and having the substituents and structural features described herein. Also described are pharmaceutical compositions and medicaments that include compounds of formula I, as well as methods of using such PI3K modulators, alone and in combination with other therapeutic agents, to treat diseases or conditions that are mediated or dependent on PI3K dysregulation.

Description

Benzoxazepine compounds and methods of use thereof

Cross reference to related applications

The present non-provisional application filed pursuant to 37CFR § 1.53(b) claims the benefit of U.S. provisional application No. 62/188,018 filed 2015, 7, 2 and U.S. provisional application No. 62/205,127 filed 2015, 8, 14, under 35USC § 119(e), which is incorporated by reference herein in its entirety.

Technical Field

The present invention relates generally to benzoxazepines

A pharmaceutical combination of oxazolidinone compounds which have activity against hyperproliferative disorders such as cancer. The invention also relates to methods of using the compounds for in vitro, in situ, and in vivo diagnosis or treatment of mammalian cells or associated pathological conditions.

Background

Upregulation of phosphoinositide-3-kinase (PI3K)/Akt signaling pathways is a common feature of most cancers (Yuan and Cantley (2008) Oncogene 27: 5497-510). genetic variations of the pathways have been detected in a variety of human cancers (Osaka et al (2004) Apoptosis 9:667-76) and act primarily to stimulate cell proliferation, migration and survival. activation of the pathways occurs after point mutation or expansion of the PIK3CA gene that activates the p110 (alpha) PI3K isoform (Hennessy et al (2005) nat. Rev. Drug Discov.4: 988-1004). tumor suppressor PTEN (a phosphatase with opposite function to PI3K) genetic deletion or loss of functional mutation in the PTEN (a phosphatase with opposite function to PI3K) also increases PI3K pathway signaling (Zhang and Yin (Clin) Resr.447. Res.2010: 25: 16: tret 16: 25. as markers for increased downstream of Cancer kinase activity such as Ak-5-kinase activity (Ak et al) has been proposed as a downstream of Cancer kinase-35-kinase (Ak et al).

Phosphatidylinositol-3-kinase (PI3K) is the major signaling node for critical survival and growth signals of lymphomas and is antagonized by the activity of the phosphatase PTEN the phosphoinositide 3-dependent kinase (PI3K) signaling pathway is the most severely deregulated pathway in hormone receptor positive breast cancer (HR + BC) the PI3K pathway is deregulated in aggressive forms of lymphomas (abbaker (2007) L eukemia 21:2368 and 2370), 8% of D L BC L (diffuse large B-cell lymphoma) cancers have missense mutations in PI3CA (phosphatidylinositol-3-kinase catalytic subunit α) and are 37% PTEN negative by immunohistochemical testing.

Phosphatidylinositol is one of a variety of phospholipids found in cell membranes and is involved in intracellular signal transduction. Cell signaling via 3' -phosphorylated phosphoinositides has been implicated in a variety of cellular processes such as malignant transformation, growth factor signaling, inflammation and immunity (Rameh et al (1999) J.biol chem. 274: 8347-8350). The enzyme responsible for the production of these phosphorylated signaling products, phosphatidylinositol-3-kinase (also known as PI3 kinase or PI3K), was originally identified as having activity associated with viral oncoproteins and growth factor receptor tyrosine kinases, which phosphorylate Phosphatidylinositol (PI) and its phosphorylated derivatives at the 3' -hydroxyl of the inositol ring (Panayotou et al (1992) Trends Cell Biol 2: 358-60). Phosphoinositide-3-kinase (PI3K) is a lipid kinase that phosphorylates lipids at the 3-hydroxyl group of the inositol ring (Whitman et al (1988) Nature,332: 664). The 3-phosphorylated phospholipids generated by PI3 kinase (PIP3) act as second messengers that recruit kinases such as Akt and PDK1 (phosphoinositide-dependent kinase 1) having a lipid binding domain including the Pleckstrin Homology (PH) region (Vivanco et al (2002) Nature Rev. Cancer 2: 489; Phillips et al (1998) Cancer 83: 41).

The PI3 kinase family includes at least 15 different enzymes subdivided by structural homology and classified into three classes according to sequence homology and products formed by enzyme catalysis, the class I PI3 kinase is composed of 2 subunits, 110kd catalytic subunit and 85kd regulatory subunit, which contains the SH2 domain and binds to tyrosine residues phosphorylated by growth factor receptors or oncogene products having tyrosine kinase activity, thereby inducing PI3K activity of the p110 catalytic subunit, which phosphorylates its lipid substrates, the class I PI3 kinase involves important signal transduction events downstream of cytokines, integrins, growth factors and immunoreceptors, which suggests that controlling this pathway may lead to important therapeutic effects such as regulation of cell proliferation and canceration, the class I PI3K phosphorylates Phosphatidylinositol (PI), phosphatidylinositol-4-phosphate and phosphatidylinositol-4, 5-diphosphate (5634) to produce phosphatidylinositol-3-phosphate (PIP), phosphatidylinositol-3, 4-diphosphate and phosphatidylinositol-3, 4-diphosphate, and phosphatidylinositol-K, respectively, and the class II phosphatidylinositol-K phosphorylates are repeated in the class I PI 3-III phosphorylation of phosphatidylinositol-3 genes, such as PI-3526Variants show that the key PI3 Kinase isoform in Cancer is class I PI3 Kinase P110 α (Samuels et al (2004) Science 304: 554; US 5824492; US 5846824; US 6274327.) other isoforms may be important in Cancer and are also involved in cardiovascular and immunoinflammatory diseases (Workman P (2004) Biochem Soc Trans 32:393 396; Patel et al (2004) Proc. am. Absoc. of Cancer Res. (Abstract L B-247)95th Annulaleting, March 27-31, Ordo, Florida, USA; Ahmadi K and Wateld MD (2004) "osteophositide 3-Kinase: functions and memescenses" Encyclopedia of biochemistry chemistry (L nai W2, L) frequently found in head and neck cancers, ovarian cancers of liver, lung hormone 6740, ovarian hormone 110 α, lung hormone- α, colon receptor for colon Cancer⁺) Breast cancer tumors have the PIK3CA mutation. PTEN abnormalities have been found in glioblastoma, melanoma, prostate cancer, endometrial cancer, ovarian cancer, breast cancer, lung cancer, head and neck cancer, hepatocellular cancer, and thyroid cancer.

PI3 kinase (PI3K) is a heterodimer composed of p85 and p110 subunits (Otsu et al (1991) Cell 65: 91-104; Hiles et al (1992) Cell 70: 419-29). four different class I PI3K, designated PI3K α, β, and ω and each composed of a different 110kDa catalytic subunit and a regulatory subunit, three of the catalytic subunits, p110 α, p110 β, and p110, each interact with the same regulatory subunit p85, while p110 γ interacts with a different regulatory subunit p 101. these PI3K each differ in the expression pattern in human cells and tissues. in each of PI3K α, PI3 β and subtypes, the p85 subunit interacts via its SH2 domain with the phosphorylated tyrosine residues in the target protein (present in a suitable sequence background) to localize the PI3 kinase to the plasma membrane (Rameh et al (1995, Cell 83: 821: 789-1992 et al; Vocoi 9: Vocai-93).

The PI3 kinase/Akt/PTEN pathway is an attractive target for cancer drug development because such drugs are expected to inhibit cell proliferation, inhibit signaling from mesenchymal cells that maintain cancer cell survival and chemoresistance, reverse the repression of apoptosis, and overcome cancer cells' intrinsic resistance to cytotoxic agents. PI3K is activated by receptor tyrosine kinase signaling and by mutations in the p110 catalytic subunit of activating PI3K, loss of the tumor suppressor PTEN or by rare activating mutations in AKT.

Taselisib (GDC-0032, Roche RG7604, CAS registry No.1282512-48-4, Genentech Inc.) was named 2- (4- (2- (1-isopropyl-3-methyl-1H-1, 2, 4-triazol-5-yl) -5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

9-yl) -1H-pyrazol-1-yl) -2-methylpropanamide, having potent PI3K activity (WO 2011/036280; US 8242104; US8343955) and being studied in patients with locally advanced or metastatic solid tumors Taselisib (GDC-0032) is a β -isoform retention inhibitor of the PI3K catalytic subunit, is more selective for the α subunit than for the β subunit 31-fold compared to wild-type PI3K α Taselisib exhibits a higher selectivity for the mutant PI3K α isoform (oliver AG et al, AACR 2013. abstrate DDT 7-01) Taselisib is currently being developed for the treatment of patients with Estrogen Receptor (ER) positive, HER2 negative metastatic breast cancer (mBC) and non-small cell lung cancer (NSC L C) patients in phase Ia studies with single drug Taselisib 539, patients with observed response to piisib 3 et al (jspr 3. f) with observed in response to piisb 3 PR 3. PR 3b 3 et al).

Recent clinical data for PI3K inhibitors have suggested PI3K activity as a source of gastrointestinal toxicity (Akinley et al phosphatogenic linosol 3-kinase (PI3K) inhibitors as Cancer therapeutics "Journal of Hematology & Oncology 2013,6: 88-104; C.Saura et al" Phase Ib Study of the PI3K Inhibitor Taselisib (GDC-0032) in Combination with L of enzyme in Patientsw Hostime Receptor Reception-Positive Advanced Cancer "San Annonio Brenstarch Symposium-December 12,2014(PD 5-2; L optiez et al" tactisib a selection, a of PIGIy 3, PIGIy assay K3 and Oncology 3 and Oncology 201426).

The reaction mixture of Idelalisib (GS-1101, CA L-101,

gilead Sciences Inc., CASreg.No.870281-82-6, 5-fluoro-3-phenyl-2- [ (1S) -1- (7H-purin-6-ylamino) propyl]-4(3H) -quinazolinone) is a selective PI3K (delta) inhibitor and is approved for the treatment of chronic lymphocytic leukemia (C LL) (US 6800620; US 6949535; US 8138195; US 8492389; US 8637533; US 8865730; US 8980901; RE 44599; RE 44638). diarrhea and colitis are among the most common adverse events reported after Idelalisib treatment (Brown et al "Idelalisib, an inhibitor of phosphatodynolosol 3-kinase 110d, for replayed/recovered chronic lymphocytic leukemia 2014123 (22): 3390-3397;

Prescribing Information 2014；

REMS FactSheet). The significant GI toxicity observed following treatment with idelalisib is consistent with the hypothesis that inhibition of PI3K (delta) is the source of gastrointestinal toxicity.

There is a need for additional modulators of PI3K α for the treatment of cancer, in particular inhibitors of PI3K α that are selective for tumors expressing mutant PI3K α over cells expressing non-mutant PI3K α there is a particular need for agents that selectively inhibit the PI3K α isoform over PI3K β, PI3K, and PI3K γ isoforms, which can be expected to result in an enhanced therapeutic window.

Disclosure of Invention

The present invention relates generally to benzoxazepines having selective activity in modulating mutant forms of the PI3K α (alpha) isoform and having the structure of formula I

Oxazolidinone compounds:

and stereoisomers, geometric isomers, tautomers and pharmaceutically acceptable salts thereof. Various substituents are defined herein.

Another aspect of the invention is a pharmaceutical composition comprising a benzoxazepine of formula I

An oxazolidinone compound and a pharmaceutically acceptable carrier, glidant, diluent, or excipient.

Another aspect of the invention is a method of treating cancer in a patient having cancer comprising administering to the patient a therapeutically effective amount of a benzoxazepine of formula I

An oxazolidinone compound.

Another aspect of the invention is a kit for the therapeutic treatment of breast cancer, comprising:

a) benzoxazepines of the formula I

An oxazolidinone compound; and

b) instructions for therapeutic treatment of breast cancer.

Drawings

FIGS. 1A and 1B show the x-ray cocrystal structure of PI3K α (alpha) with A) taselisib (GDC-0032), and B) Compound 106.

Figures 2A and 2B show the inhibition of KP L4 tumor growth after treatment with a) compound 102 and B) compound 103.

Detailed Description

Reference will now be made in detail to certain embodiments of the invention, examples of which are illustrated in the accompanying drawings and formula. While the invention will be described in conjunction with the listed embodiments, it will be understood that they are not intended to limit the invention to these embodiments. On the contrary, the invention is intended to cover all alternatives, modifications and equivalents, which may be included within the scope of the invention as defined by the appended claims. Those skilled in the art will recognize a variety of methods and materials similar or equivalent to those described herein that can be used in the practice of the present invention. The present invention is in no way limited to the methods and materials described. If one or more of the incorporated documents, patents, and similar materials differ or contradict the present application (including but not limited to defined terms, usage of terms, described techniques, etc.), the present application controls. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned in this application are incorporated in their entirety by reference into this application. Unless otherwise indicated, the nomenclature used herein is based on the IUPAC systematic nomenclature

Definition of

When stating the number of substituents, the term "one or more" refers to substitution of one substituent to the maximum possible number, i.e., replacement of one hydrogen by a substituent to replacement of all hydrogens. The term "substituent" means an atom or group of atoms that replaces a hydrogen atom on a parent molecule. The term "substituted" means that the specified group bears one or more substituents. When any group can carry multiple substituents and a variety of possible substituents are provided, the substituents are independently selected and need not be the same. The term "unsubstituted" means that the specified group bears no substituents. The term "optionally substituted" means that the specified group is unsubstituted or substituted with one or more substituents independently selected from the possible substituents. When stating the number of substituents, the term "one or more" refers to substitution of one substituent to the maximum possible number, i.e., replacement of one hydrogen by a substituent to replacement of all hydrogens.

The term "alkyl" as used herein refers to a compound having 1 to 12 carbon atoms (C)₁-C₁₂) Wherein the alkyl group may be optionally independently substituted with one or moreThe following substituents. In another embodiment, the alkyl group has 1 to 8 carbon atoms (C)₁-C₈) Or 1-6 carbon atoms (C)₁-C₆). Examples of alkyl groups include, but are not limited to, methyl (Me, -CH)₃) Ethyl (Et-CH)₂CH₃) 1-propyl (n-Pr, n-propyl, -CH)₂CH₂CH₃) 2-propyl (i-Pr, isopropyl, -CH (CH)₃)₂) 1-butyl (n-Bu, n-butyl, -CH)₂CH₂CH₂CH₃) 2-methyl-1-propyl (i-Bu, isobutyl, -CH)₂CH(CH₃)₂) 2-butyl (s-Bu, sec-butyl, -CH (CH)₃)CH₂CH₃) 2-methyl-2-propyl (t-Bu, tert-butyl, -C (CH)₃)₃) 1-pentyl (n-pentyl, -CH)₂CH₂CH₂CH₂CH₃) 2-pentyl (-CH (CH)₃)CH₂CH₂CH₃) 3-pentyl (-CH (CH)₂CH₃)₂) 2-methyl-2-butyl (-C (CH)₃)₂CH₂CH₃) 3-methyl-2-butyl (-CH (CH)₃)CH(CH₃)₂) 3-methyl-1-butyl (-CH)₂CH₂CH(CH₃)₂) 2-methyl-1-butyl (-CH)₂CH(CH₃)CH₂CH₃) 1-hexyl (-CH)₂CH₂CH₂CH₂CH₂CH₃) 2-hexyl (-CH (CH)₃)CH₂CH₂CH₂CH₃) 3-hexyl (-CH (CH)₂CH₃)(CH₂CH₂CH₃) 2-methyl-2-pentyl (-C (CH))₃)₂CH₂CH₂CH₃) 3-methyl-2-pentyl (-CH (CH)₃)CH(CH₃)CH₂CH₃) 4-methyl-2-pentyl (-CH (CH)₃)CH₂CH(CH₃)₂) 3-methyl-3-pentyl (-C (CH)₃)(CH₂CH₃)₂) 2-methyl-3-pentyl (-CH (CH)₂CH₃)CH(CH₃)₂) 2, 3-dimethyl-2-butyl (-C (CH)₃)₂CH(CH₃)₂)3, 3-twoMethyl-2-butyl (-CH (CH)₃)C(CH₃)₃1-heptyl, 1-octyl, and the like.

The terms "carbocycle", "carbocyclyl", "carbocycle", and cycloalkyl "refer to a compound having 3 to 12 carbon atoms (C)₃-C₁₂) Monovalent non-aromatic saturated or partially unsaturated rings in the form of a single ring or in the form of a ring having 7 to 12 carbon atoms. Bicyclic carbocycles having 7 to 12 atoms may be arranged, for example, as bicyclo [4,5 ]]System, bicyclo [5,5 ]]System, bicyclo [5,6 ]]Systems or bicyclo [6,6 ]]The bicyclic carbocyclic ring having 9 or 10 ring atoms of the system may be arranged as a bicyclo [5,6 ]]Systems or bicyclo [6,6 ]]Systems or arrangements for bridging systems such as bicyclo [2.2.1]Heptane, bicyclo [2.2.2]Octane and bicyclo [3.2.2]Nonane. Spiro carbocyclyl moieties are also included within the scope of this definition. Examples of spiro carbocyclyl moieties include [2.2]Pentyl, [2.3 ] or]Hexyl and [2.4 ]]A heptyl group. Examples of monocyclic carbocycles include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, 1-cyclopent-1-enyl, 1-cyclopent-2-enyl, 1-cyclopent-3-enyl, cyclohexyl, 1-cyclohex-1-enyl, 1-cyclohex-2-enyl, 1-cyclohex-3-enyl, cyclohexadienyl, cycloheptyl, cyclooctyl, cyclononyl, cyclodecyl, cycloundecyl, cyclododecyl, and the like. Carbocyclyl is optionally independently substituted with one or more substituents described herein.

"aryl" means having 6 to 20 carbon atoms (C)₆-C₂₀) A monovalent aromatic hydrocarbon group of (a), which is obtained by: one hydrogen atom is removed from a single carbon atom in the parent aromatic ring system. Some aryl groups are represented in the exemplary structures as "Ar". Aryl includes bicyclic groups containing an aromatic ring fused to a saturated, partially unsaturated, or aromatic carbocyclic ring. Typical aryl groups include, but are not limited to, groups derived from benzene (phenyl), substituted benzenes, naphthalenes, anthracenes, biphenyls, indenyls, indanyl, 1, 2-dihydronaphthalene, 1,2,3, 4-tetrahydronaphthalene, and the like. Aryl groups may be optionally substituted independently with one or more substituents described herein.

The terms "heterocycle (heterocyclic)", "heterocyclyl (heterocyclic)" and "heterocyclic" are used interchangeably herein and are meant to haveA saturated or partially unsaturated (i.e., having one or more double and/or triple bonds in the ring) carbocyclic group of 3 to about 20 ring atoms in which at least one ring atom is a heteroatom selected from nitrogen, oxygen, phosphorus, and sulfur, the remaining ring atoms being C, wherein one or more ring atoms are optionally independently substituted with one or more substituents described below. The heterocycle may be a monocyclic ring having 3 to 7 ring members (2 to 6 carbon atoms and 1 to 4 heteroatoms selected from N, O, P and S) or a bicyclic ring (e.g., bicyclo [4,5 ] having 7 to 10 ring members (4 to 9 carbon atoms and 1 to 6 heteroatoms selected from N, O, P and S)]System, bicyclo [5,5 ]]System, bicyclo [5,6 ]]Systems or bicyclo [6,6 ]]Systems) heterocycles are described in Paquette, L eo A., "Principles of Modern Heterocyclic Chemistry" (W.A. Benjamin, New York, 1968) (in particular chapters 1,3, 4,6, 7 and 9), "The Chemistry of Heterocyclic Compounds, A series of monograms" (John Wiley&Sons, New York,1950to present) (especially volumes 13, 14, 16, 19 and 28); and J.am.chem.Soc. (1960) 82: 5566. "Heterocyclyl" also includes groups in which a heterocyclyl group is fused to a saturated ring, a partially unsaturated ring, an aromatic carbocyclic ring, or an aromatic heterocyclic ring. Examples of heterocycles include, but are not limited to, morpholin-4-yl, piperidin-1-yl, piperazinyl, piperazin-4-yl-2-one, piperazin-4-yl-3-one, pyrrolidin-1-yl, thiomorpholin-4-yl, S-dioxothiomorpholin-4-yl, azetidin-1-yl, octahydropyrido [1,2-a ] o]Pyrazin-2-yl, [1,4 ]]Diazepan-1-yl, pyrrolidinyl, tetrahydrofuryl, dihydrofuranyl, tetrahydrothienyl, tetrahydropyranyl, dihydropyranyl, tetrahydrothiopyranyl, piperidino, morpholino, thiomorpholino, thionohexyl, piperazinyl, homopiperazinyl, azetidinyl, oxetanyl, thietanyl, homopiperidinyl, oxepanyl, thietanyl, oxazepinyl

Radical diaza

Radical, sulfur nitrogen hetero

A group, 2-pyrrolinyl group, 3-pyrrolinyl group, indolinyl group, 2H-pyranyl group, 4H-pyranyl group, dioxanyl group, 1, 3-dioxolanyl group, pyrazolinyl group, dithiacyclohexyl group, dithiocyclopentyl group, dihydropyranyl group, dihydrothienyl group, dihydrofuryl group, pyrazolidinyl group, imidazolinyl group, imidazolidinyl group, 3-azabicyclo [3.1.0 ] group]Hexyl, 3-azabicyclo [4.1.0]Heptyl, azabicyclo [2.2.2]Hexyl, 3H-indolyl, quinolizinyl and N-pyridylurea groups. Spiro heterocyclyl moieties are also included within the scope of this definition. Examples of spiro heterocyclyl moieties include azaspiro [2.5 ]]Octyl and azaspiro [2.4 ]]A heptyl group. Examples of heterocyclyl groups in which 2 ring atoms are substituted with an oxo (═ O) moiety are pyrimidinone groups and 1, 1-dioxo-thiomorpholinyl groups. The heterocyclyl is optionally independently substituted with one or more substituents described herein.

The term "heteroaryl" refers to a monovalent aromatic group in the form of a 5,6 or 7 membered ring containing one or more heteroatoms independently selected from nitrogen, oxygen and sulfur and includes fused ring systems (wherein at least one ring is aromatic) having 5 to 20 atoms. Examples of heteroaryl groups are pyridyl (including, for example, 2-hydroxypyridyl), imidazolyl, imidazopyridyl, pyrimidinyl (including, for example, 4-hydroxypyrimidinyl), pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furanyl, thienyl, isoxazolyl, thiazolyl, oxadiazolyl, oxazolyl, isothiazolyl, pyrrolyl, quinolyl, isoquinolyl, tetrahydroisoquinolinyl, indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl, indolizinyl, phthalazinyl, pyridazinyl, triazinyl, isoindolyl, pteridinyl, purinyl, oxadiazolyl, triazolyl, thiadiazolyl, furazanyl, benzofurazanyl, benzothienyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinoxalinyl, naphthyridinyl, and furopyridinyl. Heteroaryl groups are optionally independently substituted with one or more substituents described herein.

The terms "treatment" and "treatment" refer to a therapeutic treatment in which the aim is to slow down (lessen) the development or spread of an undesired physiological change or disorder, such as arthritis or cancer. For purposes of this application, beneficial or desired clinical results include, but are not limited to, alleviation of symptoms, diminishment of extent of disease, stabilized (i.e., not worsening) state of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, and remission (whether partial or total), whether detectable or undetectable. "treatment" may also refer to prolonging survival compared to survival expected in the absence of treatment. Those in need of treatment include those suffering from a condition or disorder.

The phrase "therapeutically effective amount" refers to an amount of a compound of the present invention that (i) treats a particular disease, condition, or disorder; (ii) alleviating, ameliorating or eliminating one or more symptoms of a particular disease, condition or disorder; or (iii) preventing or delaying the onset of one or more symptoms of a particular disease, condition, or disorder described herein. In the case of cancer, a therapeutically effective amount of the drug may reduce the number of cancer cells; reducing the size of the tumor; inhibit (i.e., slow to some extent and preferably stop) cancer cell infiltration into peripheral organs; inhibit (i.e., slow to some extent and preferably stop) tumor metastasis; inhibit tumor growth to some extent; and/or alleviate one or more symptoms associated with cancer to some extent. The drug may prevent the growth of cancer cells and/or kill existing cancer cells to some extent, and may be cytostatic and/or cytotoxic. For cancer therapy, efficacy can be determined, for example, by assessing time to disease progression (TTP) and/or determining Response Rate (RR).

More specific examples of such cancers include squamous cell cancer (e.g., epithelial squamous cell cancer), lung cancer, including small-cell lung cancer, non-small cell lung cancer ("NSC L C"), adenocarcinoma of the lung and squamous carcinoma of the lung, peritoneal cancer, hepatocellular cancer, gastric or stomach cancer, including gastrointestinal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatoma, breast cancer, colon cancer, rectal cancer, colorectal cancer, endometrial or uterine carcinoma, salivary gland carcinoma, renal or renal cancer, prostate cancer, vulval cancer, thyroid cancer, hepatic carcinoma, anal carcinoma, penile carcinoma, and head and neck cancer.

"hematological/oncology" is a sub-discipline under medicine in some centers, and considered a separate department in other centers (where there is also a surgical and radio oncologist), not all hematological diseases are malignant ("cancerous"), these other hematological diseases may also be treated by hematologists and/or oncologists, hematological malignancies may also be treated by hematologists, hematological malignancies may be derived from two major blood cell lineages: myeloid and lymphoid cell lineages, myeloid-like cell lineages generally produce granulocytes, erythrocytes, platelets, macrophages and mast cells, lymphoid cell lineages produce B cells, T cells, NK cells and plasma cell lineages, lymphocytic leukemias and from lymphoblastic cell lineages, and lymphoblastic cell lineages generally produce granulocytic cells, myeloid cells and erythroid cells, myelodysplastic leukemias (365954), acute myelogenous leukemias (365954), acute myelodysplastic leukemias (LL), and acute lymphoblastic leukemia (365954), and acute myelodysplastic leukemia (LL), including all four myelogenous leukemias (36594654).

A "chemotherapeutic agent" is a compound useful in the treatment of cancer, regardless of the mechanism of action. Classes of chemotherapeutic agents include, but are not limited to, alkylating agents, antimetabolites, spindle toxic plant alkaloids, cytotoxic/antitumor antibiotics, topoisomerase inhibitors, antibodies, photosensitizers, and kinase inhibitors. The chemotherapeutic agent is included inCompounds used in "targeted therapy" and conventional chemotherapy. Examples of chemotherapeutic agents include: ibrutinib (IMBRUVICA)^TMAPCI-32765, Pharmacyclics Inc./Janssen Biotech Inc.; CAS Reg.No.936563-96-1, US7514444), idelalisib (formerly CA L-101, GS 1101, GS-1101, Gilead Sciences Inc.; CASReg.No. 1146702-54-6), erlotinib (erlotinib) ((formerly CA L-101)

Genentech/OSI Pharm.), docetaxel (docetaxel)

Sanofi-Aventis), 5-FU (fluorouracil, 5-fluorouracil, CAS No.51-21-8), gemcitabine (gemcitabine) (

L illy), PD-0325901 (CASNO.391210-10-9, Pfizer), cisplatin (cissplatin) ((II)

(SP-4-2) -diamine platinum (II) dichloride, cis-diamine platinum (II) dichloride, CAS No.15663-27-1), carboplatin (CAS No.41575-94-4), paclitaxel (paclitaxel) ((II)

Bristol-Myers Squibb Oncology, Princeton, N.J.), trastuzumab (trastuzumab)

Genentech), temozolomide (temozolomide) (4-methyl-5-oxo-2, 3,4,6, 8-pentaazabicyclo [4.3.0 ]]Nonane-2, 7, 9-triene-9-carboxamide, CAS No.85622-93-1,

schering Plough), tamoxifen (tamoxifen) ((Z) -2- [4- (1, 2-diphenylbut-1-enyl)) Phenoxy radical]-N, N-dimethylethylamine,

) Doxorubicin (doxorubicin) ((d))

CAS No.2321

-92-8), Akti-1/2, HPPD and rapamycin (rapamycin).

Chemotherapeutic agents include inhibitors of B cell receptor targets, such as BTK, Bcl-2, and JAK inhibitors.

Further examples of chemotherapeutic agents include oxaliplatin (oxaliplatin) ((a))

Sanofi), bortezomib (bortezomib), (b), (d), (

Millennium Pharm.), sunitinib (sutent), (a mixture of them)

SU11248, Pfizer), letrozole (letrozole), (L-Toxole)

Novartis), imatinib mesylate (imatinib mesylate), (I) and (II) a pharmaceutically acceptable salt thereof

Novartis), X L-518 (Mek inhibitor, Exelixis, WO 2007/044515), ARRY-886(Mek inhibitor, AZD6244, Array BioPharma, Astra Zeneca), SF-1126(PI3K inhibitor, Semaform Pharmaceuticals), BEZ-235(PI3K inhibitor, Novartis), X L-147 (PI3K inhibitor, Exelixis), PTK787/ZK 584(Novartis), fulvestrant (fulvestrant) ((fulvestrant)

AstraZeneca), leucovorin (folinic)acid), rapamycin (sirolimus),

wyeth), lapatinib (lapatinib)

GSK572016 (Glaxo Smith Kline), Lonafanib (Lonafarnib) (SARASAR)^TMSCH 66336, Schering Plough), Sorafenib (sorafenib) ((Schering Plough)

BAY43-9006, Bayer L abs), gefitinib (gefitinib) ((B)

AstraZeneca), irinotecan (irinotecan), (

CPT-11, Pfizer), tipifarnib (ZARNESTRA)^TM,Johnson&Johnson)、 ABRAXANE^TM(without Cremophor), albumin engineered nanoparticle formulations of paclitaxel (American pharmaceutical Partners, Schaumberg, Il), vandetanib (rINN, ZD6474,

AstraZeneca), chlorambucil (chlorambucil), AG1478, AG1571(SU 5271; sugen), temsirolimus (temsirolimus) ((II)

Wyeth), pazopanib (pazopanib) (GlaxoSmithKline), canfosfamide ((Wyeth)

Telik), thiotepa and cyclophosphamide (cyclophosphamide) ((Telik)

) (ii) a Alkyl sulfonates such as busulfan, improsulfan, and piposulfan; aziridines such as benzotepa (benzodopa), carboquone (carboquone), metotepipa (meturedpa), and uredepa (uredpa); ethyleneimine (ethylenimine) and methylaminoacridine (methylamylamine) groups including hexamethylmelamine (altretamine), triimizine (triethyleneamine), triethylenephosphoramide (triethylenephosphoramide), and trimethymemimine (trimethymemimine); annonaceous acetogenins (especially bullatacin and bullatacin); camptothecin (including the synthetic analog topotecan); bryostatin; callystatin; CC-1065 (including its adozelesin (adozelesin), carvelesin (carzelesin), and bizelesin (bizelesin) synthetic analogs); cryptophycins (especially cryptophycin 1 and cryptophycin 8); dolastatin (dolastatin); duocarmycins (including the synthetic analogs KW-2189 and CB1-TM 1); eiscosahol (eleutherobin); pancratistatin; sarcodictyin; spongistatin; nitrogen mustards such as chlorambucil (chlorambucil), chlorambucil (chlorenaphazine), chlorophosphamide (chlorophosphamide), estramustine (estramustine), ifosfamide (ifosfamide), mechlorethamine (mechlorethamine), mechlorethamine hydrochloride (mechlorethamine oxide hydrochloride), melphalan (melphalan), neomustard (novembichin), benzene mustard cholesterol (phenyleneterester), prednimustine (prednimustine), trofosfamide (trofosfamide) and uramustine (uracil mustard); nitroureas such as carmustine (carmustine), chlorouretocin (chlorozotocin), fotemustine (fotemustine), lomustine (lomustine), nimustine (nimustine) and ramustine (ranirnustine); antibiotics, such as enediyne (enediyne) antibiotics (e.g., calicheamicin, including calicheamicin γ 1I and calicheamicin ω I1(Angew chem. intel. ed. engl. (1994)33: 183-186); dynemicin, including dynemicin A; bisphosphonates, such as clodronate; esperamicin (esperam;)icins and neooncostatin chromophore (neocarzinostatin chromophore) and related chromoproteins enediyne antibiotics chromophore (related chromoprotein endinecrotizing chromozophore), aclacinomycin, actinomycin (actinomycin), aureomycin, azaserine (azaserine), bleomycin (bleomycin), actinomycin C (cactinomycin), carabicacin, carminomycin (carmycin), carzinomycin, carminomycin (carmycin), paramycin (carmustine), oxytocin (streptomycin), daunorubicin, dirithromycin (daptomycin), dirithromycin D (dactinomycin), daunorubicin (daunorubicin), norubicin (paradoxycycline), norubicin (gentamycin), norubicin, doxycycline, adriamycin (gentamycin), doxycycline (gentamycin), doxycycline (gentin, doxycycline), doxycycline (gentin), doxycycline (gentin, doxycycline), doxycycline (doxycycline), doxycycline (doxycycline, doxycyclineterone), methyl androsterone propionate (dromostanolone propionate), epithioandrostanol (epitiostanol), mepiquat chloride (mepiquitazone), and testolactone (testolactone); anti-adrenergic agents (anti-adrenals), such as aminoglutethimide, mitotane and trilostane; folic acid supplements such as folinic acid (frilic acid); acetoglucurolactone (acegultone); an aldophosphamide glycoside (aldophosphamideglycoside); aminolevulinic acid (aminolevulinic acid); eniluracil (eniluracil); amsacrine (amsacrine); bestrabuucil; bisantrene; idazot (edatraxate); desphosphamide (defofamine); colchicine (demecolcine); diazaquinone (diaziqutone); elfornitine; ammonium etitanium acetate; epothilone (epothilone); etoglut (etoglucid); gallium nitrate (gallimnitrate); hydroxyurea (hydroxyurea); lentinan (lentinan); lonidamine (lonidainine); maytansinol (maytansinoid) classes such as maytansinoid (maytansine) and ansamitocins (ansamitocins); mitoguazone (mitoguzone); mitoxantrone (mitoxantrone); mopidanmol; rhizobia (nitrarine); pentostatin (pentostatin); methionine mustard (phenamett); pirarubicin (pirarubicin); losoxantrone (losoxantrone); podophyllinic acid (podophyllic acid); 2-ethylhydrazine (2-ethylhydrazine); procarbazine (procarbazine);

polysaccharide complex (A), (B)

polysaccharide complex) (JHS natural products, Eugene, OR); razoxane (rizoxane); rhizomycin (rhizoxin); sizofuran (sizofiran); germanium spiroamines (spirogyranium); tenuazonic acid (tenuazonic acid); triimine quinone (triaziquone); 2,2 ', 2 "-trichlorotriethylamine (2, 2', 2" -trichlorotriethylamine); trichothecenes (trichothecenes) (especially T-2 toxin, veracurin A, bacillocin A (roridin A) and anguidine); urethane (urethan); vindesine (vindesine); dacarbazine (dacarbazine); mannomustine (mannomustine); dibromomannitol (mitobronitol); IIDulcitol bromide (mitolactol); pipobromane (pipobroman); a polycytidysine; cytarabine (arabine) ("Ara-C"); cyclophosphamide; thiotepa; 6-thioguanine (6-thioguanine); mercaptopurine (mercaptoprine); methotrexate; platinum analogs such as cisplatin and carboplatin; vinblastine (vinblastine); etoposide (VP-16); ifosfamide; mitoxantrone (mitoxantrone); vincristine (vincristine); vinorelbine (vinorelbine)

Norfloxacin (novantrone); teniposide (teniposide); idazocide (edatrexate); daunorubicin (daunomycin); aminopterin (aminopterin); capecitabine (capecitabine) (capecitabine)

Roche); ibandronate (ibandronate); CPT-11; topoisomerase inhibitor RFS 2000; difluoromethylornithine (difluoromethylornithine, DMFO); retinoids (retinoids), such as retinoic acid (retinic acid); and pharmaceutically acceptable salts, acids and derivatives of any of the foregoing.

The following are also included in the definition of "chemotherapeutic agents": (i) anti-hormonal agents such as anti-estrogens (anti-estrogen) and Selective Estrogen Receptor Modulators (SERMs) including, for example, tamoxifen (including

Tamoxifen citrate), raloxifene (raloxifene), droloxifene (droloxifene), 4-hydroxytamoxifen (4-hydroxytamoxifen), trioxifene (trioxifene), raloxifene (ketoxifene), L Y117018, onapristone (onapristone), and

(toremifene citrate) and selective estrogen receptor modulators (SERDs) such as fulvestrant (fulvestrant) ((R))

Astra Zeneca); (ii) aromatase inhibitors which inhibit aromatase (aromatase regulates the production of estrogen in the adrenal gland), such as 4(5) -imidazoles, aminoglutethimide,

(megestrol acetate)) (megestrol acetate),

(exemestane; Pfizer), formestanie, fadrozole,

(vorozole) and (C) a salt thereof,

(letrozole; Novartis) and

(anastrozole; AstraZeneca), (iii) anti-androgens (anti-androgen) such as flutamide, nilutamide, bicalutamide, leuprolide and goserelin (goserelin), and troxacitabine (1,3-dioxolane nucleoside cytosine analogue) (1, 3-dioxanone nucleoside analog), (iv) protein kinase inhibitors such as MEK inhibitors such as cobicisib (WO 2007/044515), (v) lipid kinase inhibitors such as tacrolisin (GDC-0032, Genentech Inc.) (vi) antisense oligonucleotides, particularly antisense oligonucleotides involved in the inhibition of gene expression in abnormal cell proliferation, such as those of the Ras- α, such as Ras-32H, Ras-32, Ras-androgen, and (g) antisense nucleic acid kinase inhibitors such as the enzyme kinase inhibitors such as the Coxamitinib (WO 2007/044515)

Genta Inc.); (vii) ribozymes, such as VEGF expression inhibitors (e.g.

) And inhibitors of HER2 expression; (viii) vaccines, such as gene therapy vaccines, e.g.

And

rI L-2, topoisomerase 1 inhibitors such as

And

rmRH; (ix) anti-angiogenic agents, such as bevacizumab (bevacizumab) ((r))

Genentech); and pharmaceutically acceptable salts, acids and derivatives of any of the foregoing.

Therapeutic antibodies are also included in the definition of "chemotherapeutic agents", such as alemtuzumab (Campath), bevacizumab (bevacizumab) ((r))

Genentech); cetuximab (cetuximab) (C)

Imclone); palimumab (panitumumab) (panitumumab)

Amgen), rituximab (rituximab), (b)

Genentech/Biogen Idec), pertuzumab (PERTUzumab) (PERJETA)^TM2C4, Genentech), trastuzumab (trastuzumab) ((R) 2C4, Genentech)

Genentech), trastuzumab-maytansine conjugate (trastuzumab emtansine), (ii) and (iii) a pharmaceutically acceptable salt thereof

Genentech Inc.) and tositumomab (tositumomab) (BEXXAR, Corixia).

A "metabolite" is a product produced by the in vivo metabolism of a particular compound or salt thereof. Metabolites of compounds can be identified using conventional techniques known in the art and their activity can be determined using the assays described herein. Such products may result, for example, from oxidation, reduction, hydrolysis, amidation, deamidation, esterification, deesterification, enzymatic cleavage, etc., of the administered compound. Accordingly, the present invention includes metabolites of the compounds of the present invention, including compounds produced by a method comprising contacting a compound of formula I of the present invention with a mammal for a period of time sufficient to produce a metabolite thereof.

The term "package insert" refers to instructions typically contained in commercial packages of therapeutic products that contain information regarding indications, usage, dosage, administration, contraindications, and/or precautions related to the use of the therapeutic products described above.

The term "chiral" refers to a molecule that has a mirror image partner (mirror image partner) non-superimposability, while the term "achiral" refers to a molecule that can be superimposed with its mirror image partner.

The term "stereoisomers" refers to compounds having the same chemical composition but differing in the spatial arrangement of atoms or groups.

"diastereomer" refers to a stereoisomer having two or more chiral centers and whose molecules are not mirror images of each other. Diastereomers have different physical properties, such as melting points, boiling points, spectral properties, and reactivities. Mixtures of diastereomers may be separated by high resolution analytical procedures such as electrophoresis and chromatography.

"enantiomer" refers to two stereoisomers of a compound that are non-superimposable mirror images of each other.

The stereochemical definitions and common knowledge used herein generally correspond to the Chemical definitions of S.P. Parker, Ed., McGraw-HillDirectionbearing of Chemical terminms (1984) McGraw-Hill Book Company, New York; and Eliel, E.and Wilen, S., "Stereochemistry of Organic Compounds", John Wiley & Sons, Inc., New York,1994 the Compounds of the present invention may contain an asymmetric center or a chiral center and thus exist in different stereoisomeric forms the present invention is intended to include all stereoisomeric forms of the Compounds of the present invention, including but not limited to diastereomers, enantiomers and atropisomers (atropisomers) and mixtures thereof such as racemic mixtures, forming part of the present invention.

The term "tautomer" or "tautomeric form" refers to structural isomers having different energies that can interconvert through a low energy barrier. For example, proton tautomers (also referred to as proton transfer tautomers) include interconversions by migration of protons, such as keto-enol isomerization and imine-enamine isomerization. Valence tautomers include interconversions by recombination of some of the bonding electrons.

The term "pharmaceutically acceptable salt" refers to salts that are biologically or otherwise undesirable. Pharmaceutically acceptable salts include acid and base addition salts. The phrase "pharmaceutically acceptable" means that the substance or composition must be chemically and/or toxicologically compatible with the other ingredients comprising the formulation and/or the mammal being treated therewith.

The term "pharmaceutically acceptable acid addition salts" refers to those pharmaceutically acceptable salts with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, carbonic acid, phosphoric acid and organic acids selected from aliphatic, alicyclic, aromatic, aryl-aliphatic, heterocyclic, carboxylic and sulfonic organic acids such as formic acid, acetic acid, propionic acid, glycolic acid, gluconic acid, lactic acid, pyruvic acid, oxalic acid, malic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, aspartic acid, ascorbic acid, glutamic acid, anthranilic acid, benzoic acid, cinnamic acid, mandelic acid, pamoic acid, phenylacetic acid, methanesulfonic acid (methanesulfonic acid) "methanesulfonic acid (mesylate)", ethanesulfonic acid, p-toluenesulfonic acid and salicylic acid.

The term "pharmaceutically acceptable base addition salts" refers to pharmaceutically acceptable salts formed with organic or inorganic bases. Examples of acceptable inorganic bases include sodium, potassium, ammonium, calcium, magnesium, iron, zinc, copper, manganese and aluminum salts. Salts derived from pharmaceutically acceptable organic non-toxic bases include salts of: primary, secondary and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, 2-diethylaminoethanol, trimethylamine, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline, betaine, ethylenediamine, glucosamine, methylglucamine, theobromine, purine, piperazine, piperidine, N-ethylpiperidine and polyamine resins.

"solvate" refers to an association or complex of one or more solvent molecules with a compound of the invention. Examples of solvate-forming solvents include, but are not limited to, water, isopropanol, ethanol, methanol, DMSO, ethyl acetate, acetic acid, and ethanolamine.

The term "EC₅₀"is the half maximal effective concentration and means the plasma concentration of the particular compound required to achieve 50% of the maximal in vivo specific effect.

The term "Ki" is an inhibition constant and denotes the absolute binding affinity of a particular inhibitor to a receptor. If no competing ligand (e.g., radioligand) is present, it is measured using a competitive binding assay and is equal to the concentration at which the particular inhibitor occupies 50% of the receptor. Ki values can be logarithmically converted to pKi values (-log Ki), with higher values indicating exponentially greater potency.

The term "IC₅₀"is the half maximal inhibitory concentration and refers to the concentration of a particular compound required to obtain 50% inhibition of a biological process in vitro. Can connect IC₅₀Logarithmic conversion of values to pIC₅₀Value (-log IC)₅₀) Where higher values indicate exponentially greater potency. IC (integrated circuit)₅₀The values are not absolute values but depend on experimental conditions, which can be converted into absolute inhibition constants (Ki) using, for example, the Cheng-Prusoff equation (biochem. Pharmacol. (1973) 22: 3099). Other percentage rejection parameters may be calculated, such as IC₇₀、IC₉₀And the like.

The terms "compound of the invention" and "compound of formula I" include compounds of formula I and stereoisomers, geometric isomers, tautomers, solvates, metabolites, pharmaceutically acceptable salts and prodrugs thereof.

Any formula or structure given herein, including compounds of formula I, is also intended to represent hydrates, solvates, and polymorphs of the above compounds, and mixtures thereof.

Any formulae or structures given in this application (including formulaeI compounds) are also intended to represent unlabeled as well as isotopically labeled forms of the compounds. Isotopically-labeled compounds have the structure depicted in the formulae given herein, except that one or more atoms are replaced by an atom having a selected atomic mass or mass number. Examples of isotopes that can be incorporated into compounds of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, fluorine and chlorine, such as, but not limited to²H (deuterium, D),³H (tritium),¹¹C、¹³C、¹⁴C、¹⁵N、¹⁸F、³¹P、³²P、³⁵S、³⁶Cl and¹²⁵I. the invention includes various isotopically-labeled compounds of the invention, e.g., radioactive isotopes such as³H、¹³C and¹⁴those compounds of the present invention into which C is introduced. The isotopically labeled compounds described above are useful in metabolic studies, reaction kinetic studies, detection or imaging techniques such as Positron Emission Tomography (PET) or Single Photon Emission Computed Tomography (SPECT) including drug or substrate tissue distribution assays or in the treatment of patients with radioactivity. Therapeutic compounds of the invention labeled or substituted with deuterium may have improved DMPK (drug metabolism and pharmacokinetics) properties, including absorption, distribution, metabolism and excretion (ADME). Substitution with heavier isotopes such as deuterium may afford certain therapeutic advantages resulting from greater metabolic stability (e.g. increased in vivo half-life or reduced dosage requirements). Warp beam¹⁸The F-labeled compounds are useful for PET or SPECT studies. Isotopically-labeled compounds of the present invention and prodrugs thereof can generally be prepared as follows: the procedures disclosed in the schemes or examples and preparations below were performed and the non-isotopically labeled reagents were replaced with readily available isotopically labeled reagents. In addition, with heavier isotopes, especially deuterium (i.e. deuterium)²H or D) may result in some therapeutic advantages (e.g. increased in vivo half-life or reduced dosage requirements or improved therapeutic index) due to better metabolic stability. It is to be understood that deuterium in the present application is considered as a substituent in the compounds of formula (I). The concentration of the heavier isotopes, in particular deuterium, can be adjustedOver isotopic enrichment factors. Any atom in the compounds of the present invention that is not specifically designated as a particular isotope is intended to represent any stable isotope of that atom. Unless otherwise indicated, when a location is specifically designated as "H" or "hydrogen," it is understood that the location has a concentration of hydrogen that is the natural abundance isotopic composition of hydrogen. Thus, in the compounds of the present invention, any atom specifically designated as deuterium (D) is intended to represent deuterium.

Benzoxazepines

Oxazolidinone compounds

The present invention provides benzoxazepines of formula I

Oxazolidinone compounds and pharmaceutical formulations thereof, which are potentially useful for the treatment of cancer and have the following structure:

and stereoisomers, geometric isomers, tautomers and pharmaceutically acceptable salts thereof, wherein:

R¹is selected from-CH₃、-CH₂CH₃、-CH(CH₃)₂、-CHF₂、-CH₂F and-CF₃；

X is selected from:

wherein the wavy line represents the attachment site; and is

R²Selected from H, C₁-C₆Alkyl, cyclopropyl and cyclobutyl optionally substituted by F, -OCH₃or-OH.

Benzoxazepines of the formula I

Oxazolidinone compounds include those of formula Ia:

exemplary embodiments of compounds of formula Ia include those wherein R²is-CH₃And R is¹Selected from-CHF₂and-CH₂F。

Benzoxazepines of the formula I

Oxazolidinone compounds include those of formula Ib:

exemplary embodiments of compounds of formula Ib include those in which R¹Selected from-CHF₂and-CH₂F。

Exemplary embodiments of compounds of formula I include the compounds in table 1.

The compounds of formula I according to the invention may contain asymmetric or chiral centers and therefore exist in different stereoisomeric forms. All stereoisomeric forms of the compounds of the present invention, including but not limited to diastereomers, enantiomers and atropisomers and mixtures thereof, such as racemic mixtures, form part of the present invention. In some cases, stereochemistry has not been determined or has been tentatively assigned.

In addition, the present invention includes all diastereomers, including cis-trans (geometric) and conformational isomers. For example, if the compounds of formula I contain double or fused rings, then the cis and trans forms and mixtures thereof are included within the scope of the present invention.

In the structures shown herein, the compounds of the present invention are intended to include all stereoisomers if the stereochemistry of any particular chiral atom is not specified. If stereochemistry is indicated by a solid wedge or dashed line representing a particular configuration, the stereoisomer is so indicated and defined.

The compounds of the invention may exist in unsolvated forms as well as solvated forms with pharmaceutically acceptable solvents such as water, ethanol, and the like and the invention is intended to include both solvated and unsolvated forms.

The compounds of the invention may also exist in different tautomeric forms and all such forms are included within the scope of the invention. The term "tautomer" or "tautomeric form" refers to structural isomers having different energies that can interconvert through a low energy barrier. For example, proton tautomers (also referred to as proton transfer tautomers) include interconversions by proton migration, such as keto-enol isomerization and imine-enamine isomerization. Valence tautomers include interconversions by recombination of some of the bonding electrons.

Biological evaluation

The relative potency of a compound of formula I as an inhibitor of enzymatic activity (or other biological activity) can be determined as follows: the concentration at which each compound inhibits the activity to a predetermined degree is determined, and the results are compared. Generally, the preferred determination is the concentration that inhibits 50% of activity in a biochemical assay, i.e., the 50% inhibitory concentration or "IC₅₀". IC can be determined using conventional techniques known in the art₅₀The value is obtained. In general, ICs₅₀Can be determined by measuring the activity of a given enzyme in the presence of a range of concentrations of inhibitor under investigation. The experimentally obtained enzyme activity values are then plotted against the inhibitor concentration used. Concentration of inhibitor showing 50% enzyme activity (compared to activity in the absence of any inhibitor) as IC₅₀The value is obtained. Similarly, other inhibitory concentrations may be defined by appropriate activity determinations. For example, in some cases, it may be desirable to determine the 90% inhibitory concentration or IC₉₀And the like.

Exemplary compounds of formula I in table 1 were prepared, characterized, and tested for binding to PI3K in various isoforms and mutant forms according to the methods of the present invention, and have the following structure, corresponding names (ChemBioDraw, Version 12.0.2, Cambridge soft corp., Cambridge MA) and biological activity. When more than one name is associated with a compound of formula I or an intermediate, the compound should be defined in terms of chemical structure.

Table 1.

TASELISIB

The IUPAC names for the compounds designated taselisib, GDC-0032, and Roche RG7604(CAS Reg.No.1282512-48-4, Genentech Inc.): 2- (4- (2- (1-isopropyl-3-methyl-1H-1, 2, 4-triazol-5-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) -1H-pyrazol-1-yl) -2-methylpropanamide, having the structure:

including stereoisomers, geometric isomers, tautomers and pharmaceutically acceptable salts thereof.

Taselesib can be prepared and characterized as described in WO2011/036280, US8242104 and US 8343955.

pictilisib

The compounds designated pictiliib, GDC-0941, Roche, RG-7321 and pictreliib, (CAS Reg.No.957054-30-7, Genentech Inc.,) are potent multi-target class I (pan) PI3K isoform inhibitors. GDC-0941 is currently undergoing phase II clinical trials for the treatment of advanced solid tumors. GDC-0941 is named 4- (2- (1H-indazol-4-yl) -6- ((4- (methylsulfonyl) piperazin-1-yl) methyl) thieno [3,2-d ] pyrimidin-4-yl) morpholine (US 7781433; US 7750002; Folkes et al (2008) journal of Med. chem.51(18):5522-5532) and has the following structure:

including stereoisomers, geometric isomers, tautomers and pharmaceutically acceptable salts thereof.

Biochemical inhibition of PI3K isoforms

The ability of the compounds of the invention to act as PI3K α inhibitors was determined using the method of example 901, with selectivity relative to PI3K β, PI3K and PI3K γ the FP Ki data determined using the method of example 901 from US8242104 is also included in tables 2A and 2B.

Table 2A shows the biochemical inhibition of the four PI3K isoforms by the compounds of formula I of table 1, in addition, two clinically tested PI3K compounds taselisib and pictilisib are included as a comparator representative compounds of the invention exhibit strong activity on PI3K α when compared to taselisib (GDC-0032) and pictilisib (GDC-0941) and significantly enhanced selectivity over the other isoforms PI3K β, PI3K and PI3K γ in particular the selectivity ratio in the second column from the right in table 2A shows that each compound of formula I101-.

Table 2B shows biochemical inhibition of selectivity ratios by two PI3K isoforms α and PI3K α of certain comparative compounds of US8242104 and compounds with a dimethyl oxazolidin-2-one group from US 8263633 (compound 356, column 149). the comparative compounds shown in table 2B are examples from the broad genus described in each of US8242104 and US 8263633 neither US8242104 nor US 8263633 disclose compounds within the scope of the compounds of formula I of the present invention although representative comparative examples of US8242104 as described in table 2B show a PI3K α selectivity ratio >1 relative to PI3K, the maximum selectivity ratio observed is 269-fold.

Other representative examples of PI3K inhibitors, such as taselisib (WO 2011/036280; US 8242104; US8343955) and US8242104, in current clinical trials exhibit significant activity on PI3K (delta) isoforms this lack of selectivity to PI3K (delta) is consistent with the GI toxicity observed in the clinic for taselisib there is a need for PI3K α (alpha) inhibitors that contain the advantageous features of the representative example of US8242104, which lack activity on PI3K at the same time.

The unexpected nature of PI3K α selectivity is beneficial in abrogating gastrointestinal toxicity observed in clinical PI3K inhibitor candidates recent clinical data for PI3K inhibitors have implicated PI3K activity as a source of gastrointestinal toxicity (akinley et al, "pathodylinosol 3-kinase (PI3K) inhibitors as cancer therapeutics" Journal of health & Oncology 2013,6: 88-104.) see table 2 for PI3K inhibitors in clinical trials.

Since PI3K α (alpha) inhibition is significantly more selective than PI3K (delta) inhibition, the toxicity driven by compound 101-109 versus PI3K (delta) of formula I is expected to achieve a greater magnitude of clinical activity (margin) driven by PI3K α (alpha) inhibition than taselisib and piculisib tested clinically, therefore, the compounds of formula I of the present invention are useful as therapeutic agents with reduced toxicity profiles relative to drugs with greater inhibition of the normal function of PI3K β, PI3K, or PI3K γ.

TABLE 2AThe biochemical inhibitory effect of the compounds of formula I and of the comparative compounds taselisib and pictilisib on the PI3K isoform

TABLE 2BComparison of the Biochemical inhibition of PI3K isoforms of Compounds

Interaction of Compounds with PI3K

A reasonable basis for PI3K α selectivity of compounds of formula I may exist in certain binding interactions.

The ability of the compounds of the invention to specifically interact with PI3K α was determined by resolving the x-ray co-crystal structure of representative compounds having PI3K α using the method of example 902. optimal structural design of PI3K inhibitors that are selective for PI3K α isoforms relative to other isoforms may include precise positioning and arrangement of atoms and functional groups to interact with isoform-specific residues in the binding site, specifically, found in 5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

The substitution in position 2 of the ring system has a major influence on the specific activity of the compounds on PI3K α the oxazolidinone ring of the compounds of formula I is capable of multiple improved interactions with proteins relative to the triazole ring.

FIG. 1A shows the x-ray structure of taselisib, bound at the PI3K α (alpha) active site, the N4 atom of the triazole ring cannot directly bind to Tyr836 (alpha)

Distance of) or Ser774(2.74 and Ser 774)

No complementary polarity between ligand and residue fig. 1B shows the x-ray structure of compound 106 bound at the PI3K α active site and shows that the oxazolidinone ring is able to undergo multiple improved interactions with proteins relative to the triazole ring

And can form favorable polar interactions. The fluorine atom of the oxazolidinone substituent is closely contacted with the hydroxyl of Ser774

And consistent with polar interactions or non-classical hydrogen bonding (enabling favorable interactions through polarization of carbon-fluorine bonds) ((R))

Etc. "Fluorine In medicinal chemistry" (2004) ChemBiochem,5: 637-643; Zhou et al, "Fluorine Bonding-How In project-L igand Interactions" (2009) J.chem. Inf.model.,49: 2344-2355).

All compounds of the present invention contain oxazolidone rings and are capable of improved interaction with Tyr836 of PI3K α some embodiments of the present invention also contain fluorinated substituents on the oxazolidone ring and are capable of improved interaction with Ser774 of PI3K α relative to the examples of US8242104, these two binding interactions may contribute to the increased selectivity to PI3K α observed in embodiments of the present invention, residues Ser774 and Tyr836 are not unique to PI3K α isoform, PI3K contains the same residue at the same position, the enhanced isoform selectivity of the oxazolidinone inhibitor is not predicted by these crystal structures.

Oxazolidinones are structurally distinct from triazoles in that: oxazolidinones have a carbonyl group, are more polar, and are not aromatic. Triazoles have no carbonyl group, are less polar, and have aromaticity.

According to increased sp3 character and decreasedIn contrast, an increase in the ratio of sp3 carbons (# sp3 carbons/# total carbons) is associated with improved Physicochemical properties and reduced promiscuous binding, thereby reducing the risk of off-target toxicology, as described in reference L overjet et al, "Escape From fluoride", (2009) j.med.chem.,52:6752-]Imidazo [1,2-d ] s][1,4]Oxazazem

Examples of 2-position saturated heterocyclic systems of rings, US8242104 provides no teaching on methods for replacing the aromatic ring with a saturated heterocyclic ring while maintaining activity towards PI3K α.

Thus, the compounds of the invention are described in 5, 6-dihydrobenzo [ f ]]Imidazo [1,2-d ] s][1,4]Oxazazem

These optimized compounds provide significant and heretofore unknown benefits in improving molecular interactions and increasing selective activity against PI3K α, and reduced activity against PI3K the compounds of the invention are useful as therapeutic agents with an enhanced therapeutic window relative to related agents such as taselisib (GDC-0032).

Selective inhibition of mutant PI3K α (alpha)

The ability of the compounds of the invention to preferentially act against cells containing mutant PI3K α was determined by measuring inhibition of the PI3K pathway in the SW48 near isogenic cell line PI3K α wild type (parental), helical domain mutant E545K and kinase domain mutant H1047R as described in example 903.

Statistical analysis: unless otherwise stated, EC50 values represent geometric means of a minimum of 4 independent experiments. All statistics were performed using KaleidaGraph software (version 4.1.3). Student's t-test was performed using unpaired data with the same variance to compare activity against mutant and wild type cells. P <0.05 was considered significant.

Table 3A shows the inhibition of P-PRAS40 in SW48 near isogenic cells by the compounds of formula I of table 1 both exhibit increased activity on mutant PI3K α cells relative to wild-type PI3K α cells with a selectivity ratio > 2-fold and a P-value <0.05 the compounds of the invention show similar activity to taselisib, with equal or higher selectivity to taselisib in SW48 mutant PI3K α cells relative to the activity in wild-type PI3K α cells (see table 3B).

Table 3B shows that certain comparative compounds of US8242104, compounds with a dimethyloxazolidin-2-one group from US 8263633 (compound 356, column 149) and pictilisib, inhibit P-PRAS40 in SW48 near isogenic cells the comparative compounds shown in table 3B are examples from the broad genus described in each of US8242104 and US 8263633 neither US8242104 nor US 8263633 disclose compounds within the scope of the compounds of formula I of the present invention the comparative compounds contain examples of mutant PI3 cells that do not significantly increase activity relative to wild type PI3K α cells (see comparative compounds pictilisib, 375, 436, 469 and 486, for one or both tested mutations P > 0.05.) these compounds do not exhibit significantly increased activity relative to wild type PI3K α cells with respect to the comparative compounds that do not exhibit significantly increased activity in the wild type PI3 cells (see comparative compounds tasibib, 469, 46540, and for one or both tested mutations P > 0.05.) the comparative compounds that do not exhibit significantly increased activity relative to wild type PI3 cells in the wild type PI 4833 cells or the overall selectivity of the compounds taught by the test PI 678678 no compounds that provide an increase in the structure of PI equivalent to wild type PI 4253 or PI 82387 compounds in the wild type PI 82387 3 cell.

Table 3A.Inhibition of SW48 near isogenic cells by P-PRAS40 with compounds of formula I

Table 3B.Comparison of inhibition of P-PRAS40 by SW48 near isogenic cells

Antiproliferative activity of PI3K mutant tumor cells

The ability of the compounds of the invention to reduce the viability of PI3K mutant tumor cells was determined by measuring antiproliferative EC50 in HCC1954 and KP L4 cells (PI3K α mutant H1047R) and MCF7 cells (PI3K α mutant E545K) using the method of example 904, table 4 shows that compounds of formula I102, 103 and 105 inhibit the proliferation of HCC1954, KP L4 and MCF7 cells with similar levels of potency as the comparative compounds taselisib (compound 196, US8242104), piculisib and compound 436(US 8242104).

Table 4.Antiproliferative activity in mutant PI3K- α tumor cells

In vivo efficacy in tumor xenograft models

The ability of the compounds of the invention to inhibit tumor growth in an in vivo tumor xenograft model was determined using the method described in example 905 using the KP L4 breast cancer cell line (PI3K α mutant H1047R fig. 2A and 2B show that compounds 102 and 103 of formula I, respectively, are able to strongly inhibit the growth of KP L4 tumors in vivo in a dose-dependent manner using daily PO (oral) administration all doses of compounds 102 and 103 were well tolerated and no treatment-related weight loss was observed.

Administration of Compounds of formula I

The compounds of the present invention may be administered by any route suitable for the condition to be treated. Suitable routes include oral, parenteral (including subcutaneous, intramuscular, intravenous, intraarterial, intradermal, intrathecal and epidural), transdermal, rectal, nasal, topical (including buccal and sublingual), vaginal, intraperitoneal, intrapulmonary and intranasal. For local immunosuppressive therapy, the compound may be administered by intralesional administration (including perfusion or contacting the graft with an inhibitor prior to transplantation). It will be appreciated that the preferred route may vary, for example, with the condition of the recipient. When the compound is administered orally, it may be formulated into pills, capsules, tablets, and the like, together with a pharmaceutically acceptable carrier or excipient. When the compound is administered parenterally, it may be formulated with a pharmaceutically acceptable parenteral vehicle and into unit dose injectable forms, as described below.

The dose for treating a human patient may be from about 1mg to about 1000mg of a compound of formula I. A typical dose may be from about 10mg to about 300mg of the compound. The dose may be administered once daily (QID), twice daily (BID), or more frequently, depending on the pharmacokinetic and pharmacodynamic properties of the particular compound, including absorption, distribution, metabolism, and excretion. In addition, toxicity factors can affect the dosage and administration regimen. When administered orally, the pills, capsules or tablets may be taken daily or less frequently for the specified period of time. The protocol may be repeated for multiple treatment cycles.

Methods of treatment using compounds of formula I

The compounds of formula I of the present invention are useful in treating human or animal patients suffering from diseases or disorders such as cancer due to abnormal cell growth, function or behaviour associated with PI3K and may therefore be treated by a method comprising administering thereto a compound of the present application as defined above. A human or animal patient suffering from cancer may also be treated by a method comprising administering thereto a compound of the invention as defined above. The condition of the patient may thus be improved or alleviated.

The methods of the invention also include treating a cancer selected from the group consisting of breast, ovarian, cervical, prostate, testicular, genitourinary tract, esophageal, laryngeal, glioblastoma, neuroblastoma, gastric, skin, keratoacanthoma, lung, epidermoid carcinoma, large cell carcinoma, non-small cell lung carcinoma (NSC L C), small cell carcinoma, lung adenocarcinoma, bone, colon, adenoma, pancreatic, adenocarcinoma, thyroid, follicular carcinoma, undifferentiated carcinoma, papillary carcinoma, seminoma, melanoma, sarcoma, bladder carcinoma, liver carcinoma and biliary passages, kidney carcinoma, pancreatic carcinoma, myeloid disorders, lymphoma, hairy cell carcinoma, oral cavity carcinoma, nasopharyngeal carcinoma, pharyngeal cancer, lip carcinoma, tongue carcinoma, mouth carcinoma, small bowel carcinoma, colon-rectal carcinoma, large-sized carcinoma of the intestine, rectal carcinoma, brain carcinoma and central nervous system carcinoma, hodgkin's carcinoma, leukemia, bronchial carcinoma, thyroid carcinoma, intrahepatic and bile duct carcinoma, hepatocellular carcinoma, gastric carcinoma, glioma/glioblastoma, renal carcinoma, melanoma, cervical carcinoma, myeloid leukemia, chronic myeloma, leukemia, chronic myelogenous leukemia, and leukemia.

Colon, breast, cervical, gastric, lung malignancies and multiple myeloma are most likely to respond to PI3K modulators or inhibitors based on expression analysis, immunohistochemical analysis and cell line distribution.

The present invention relates to the use of a compound as described above for the treatment of cancer in a patient, wherein the cancer is selected from breast cancer and non-small cell lung cancer.

The present invention relates to the use of a compound as described above for the manufacture of a medicament for the treatment of cancer in a patient, wherein the cancer is selected from breast cancer and non-small cell lung cancer.

The present invention relates to a compound as described above for use in the treatment of cancer in a patient, wherein the cancer is selected from breast cancer and non-small cell lung cancer.

The invention as described above.

Pharmaceutical preparation

For use of the compounds of the present invention for therapeutic treatment of mammals, including humans, they are generally formulated as pharmaceutical compositions in accordance with standard pharmaceutical practice. This aspect of the invention provides a pharmaceutical composition comprising a compound of the invention and a pharmaceutically acceptable diluent or carrier.

Typical formulations are prepared by mixing a compound of the invention with a carrier, diluent or excipient.

Suitable carriers, diluents, additives and excipients are well known to those skilled in the art and include materials such as carbohydrates, waxes, water soluble and/or swellable polymers, hydrophilic or hydrophobic materials, gelatin, oils, solvents, water and the like. The particular carrier, diluent or excipient employed will depend upon the means and purpose of administering the compounds of the present invention. The solvent is generally selected based on a solvent (GRAS) deemed safe by one of skill in the art for administration to a mammal. Generally, safe solvents are non-toxic aqueous solvents such as water and other non-toxic solvents that are soluble or miscible in water. Suitable aqueous solvents include water, ethanol, propylene glycol, polyethylene glycols (e.g., PEG 400, PEG 300), and mixtures thereof. The formulations may also contain one or more buffering agents, stabilizing agents, surfactants, wetting agents, lubricating agents, emulsifying agents, suspending agents, preservatives, antioxidants, opacifiers, glidants, processing aids, colorants, sweeteners, fragrances, flavoring agents and other known additives to impart a superior appearance to the drug (i.e., a compound of the present application or a pharmaceutical composition thereof) or to aid in the manufacture of the pharmaceutical product (i.e., a pharmaceutical product).

The formulations may be prepared using conventional dissolution and mixing operations. For example, the bulk drug substance (i.e., a compound of the invention or a stabilized form of the compound (e.g., a complex with a cyclodextrin derivative or other known complexing agent)) is dissolved in a suitable solvent in the presence of one or more of the above-mentioned excipients. The compounds of the present invention are typically formulated into pharmaceutical dosage forms to provide easily controllable dosages of the drug and to enable patient compliance with prescribed regimens.

The pharmaceutical composition (or formulation) for administration may be packaged in a variety of ways depending on the method of administering the drug. Typically, the article of manufacture for dispensing comprises a container in which the pharmaceutical formulation is stored in a suitable form. Suitable containers are known to those skilled in the art and include materials such as bottles (plastic and glass), pouches, ampoules, plastic bags, metal cylinders, and the like. The container may also include anti-pry means to prevent inadvertent access to the contents of the package. Additionally, the container has a label thereon that describes the contents of the container. The tag may also include suitable warning information.

Pharmaceutical formulations of the compounds of the present application can be prepared for various routes and types of administration. For example, a compound of formula I having a desired purity may be optionally mixed with pharmaceutically acceptable diluents, carriers, excipients or stabilizers in the form of a lyophilized formulation, a milled powder or an aqueous solution (Remington's Pharmaceutical Sciences (1980) 16 th edition, Osol, a.ed.). The formulation can be carried out as follows: mixed at ambient temperature at a suitable pH and in a suitable purity with a physiologically acceptable carrier, i.e. a carrier which is non-toxic to recipients at the dosages and concentrations employed. The pH of the formulation depends primarily on the particular use and compound concentration, but can be from about 3 to about 8. Formulations in acetate buffer at pH 5 are suitable embodiments.

The drug may be stored in the form of a solid composition, a lyophilized formulation, or an aqueous solution.

The pharmaceutical compositions of the present invention will be formulated, dosed and administered in a manner consistent with good medical practice (i.e., amount, concentration, schedule, course, vehicle and route of administration). Factors to be considered in this context include the particular condition being treated, the particular mammal being treated, the clinical condition of the individual patient, the cause of the condition, the site of delivery of the drug, the method of administration, the timing of administration and other factors known to medical practitioners. The "therapeutically effective amount" of the compound to be administered will depend on the above factors considered and is the minimum amount required to ameliorate or treat the hyperproliferative disorder.

As a general proposition, the initial pharmaceutically effective amount of inhibitor per dose administered parenterally will be about 0.01-100mg/kg, i.e., about 0.1-20mg/kg, of patient body weight per day, with a typical initial range of compounds employed being 0.3 to 15 mg/kg/day.

Acceptable diluents, carriers, excipients, and stabilizers are nontoxic to recipients at the dosages and concentrations employed and include buffers such as phosphates, citrates, and other organic acids; antioxidants, including ascorbic acid and methionine; preservatives (such as octadecyl dimethyl benzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butanol or benzyl alcohol; alkyl parabens, such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrins; chelating agents, such as EDTA; sugars such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions, such as sodium; metal complexes (e.g., Zn-protein complexes); and/or nonionic surfactants, such as TWEEN^TM、PLURONICS^TMOr polyethylene glycol (PEG). The active pharmaceutical ingredient may also be embedded in microcapsules prepared, for example, by coacervation techniques or interfacial polymerization, such as hydroxymethylcellulose or gelatin-microcapsules and poly (methylmethacylate) microcapsules, respectively, in colloidal drug delivery systems (such as liposomes, albumin microspheres, microemulsions, nanoparticles and nanocapsules) or macroemulsions. See Remington's Pharmaceutical Sciences 16 th edition, Osol, A. eds (1980).

Sustained release of a compound of formula IExamples of sustained release matrices include polyesters, hydrogels (e.g., poly (2-hydroxyethyl methacrylate) or poly (vinyl alcohol)), polylactide (US3773919), L-glutamic acid and gamma-ethyl-L-glutamic acid copolymers, non-degradable ethylene-vinyl acetate, degradable lactic acid-glycolic acid copolymers such as L ron DEPOT^TM(microspheres for injection composed of lactic acid-glycolic acid copolymer and leuprolide acetate) and poly D- (-) -3-hydroxybutyric acid.

Formulations include those suitable for the routes of administration described herein. The formulations may be conveniently presented in unit dosage form and may be prepared by any of the methods known in the art of pharmacy. Techniques and formulations are generally described in Remington's pharmaceutical Sciences (Mack Publishing Co., Easton, Pa.). The above method comprises the step of bringing into association the active ingredient with the carrier as one or more accessory ingredients. In general, the formulations are prepared as follows: the active ingredient is combined with a liquid carrier or a finely divided solid carrier or both uniformly and intimately thereafter the product is shaped as required.

Formulations of a compound of formula I suitable for oral administration may be prepared as discrete units such as pills, capsules, cachets or tablets each containing a predetermined amount of a compound of formula I. Compressed tablets may be prepared as follows: the active ingredient in free-flowing form, e.g. powder or granules, optionally mixed with binders, lubricants, inert diluents, preservatives, surfactants or dispersing agents, is compressed in a suitable machine. Molded tablets may be prepared as follows: the mixture of powdered active ingredient moistened with an inert liquid diluent is moulded in a suitable machine. The tablets may optionally be coated or scored and optionally formulated for slow or controlled release of the active ingredient therefrom. Tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules such as gelatin capsules, syrups or elixirs may be prepared for oral administration. Formulations of compounds of formula I intended for oral administration may be prepared according to any method known to the art for the manufacture of pharmaceutical compositions and such compositions may contain one or more agents including sweetening agents, flavoring agents, coloring agents and preserving agents in order to provide a palatable preparation. Tablets containing the active ingredient in admixture with non-toxic physiologically acceptable excipients which are suitable for the manufacture of tablets are acceptable. These excipients may be, for example, inert diluents such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, such as corn starch or alginic acid; binding agents, such as starch, gelatin or acacia; and lubricating agents such as magnesium stearate, stearic acid or talc. Tablets may be uncoated or may be coated by known techniques including microencapsulation to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period. For example, a time delay material such as glyceryl monostearate or glyceryl distearate alone or with a wax may be employed.

For the treatment of the eye or other external tissues such as mouth and skin, the formulations may preferably be administered in the form of a topical ointment or cream containing the active ingredient in an amount of, for example, 0.075 to 20% w/w. When formulated in an ointment, the active ingredient may be used with a paraffinic or water-miscible ointment base. Alternatively, the active ingredient may be formulated as a cream together with an oil-in-water cream base. If desired, the aqueous phase of the cream base may comprise polyhydric alcohols, i.e. alcohols having two or more hydroxyl groups such as propylene glycol, butane-1, 3-diol, mannitol, sorbitol, glycerol and polyethylene glycols (including PEG 400) and mixtures thereof. Topical formulations may desirably contain compounds that enhance absorption or penetration of the active ingredient through the skin or other affected area. Examples of such skin permeation enhancers include dimethyl sulfoxide and related analogs. The oil phase of the emulsions of the present application may be constituted by known ingredients in a known manner. When the phase may comprise emulsifiers alone, it comprises, as required, a mixture of at least one emulsifier with a fat or an oil or with both a fat and an oil. Preferably, a hydrophilic emulsifier is included as a stabilizer together with a lipophilic emulsifier. It is also preferred to include both oil and fat. At the same time, the emulsifier, with or without stabilizer, constitutes the so-called milkThe wax is melted and, together with the oil and fat, constitutes the so-called emulsified cream base, which forms the oily dispersed phase of the cream. Emulsifiers and emulsion stabilizers suitable for use in the formulations herein include

60、

80. Cetostearyl alcohol, benzyl alcohol, myristyl alcohol, glyceryl monostearate and sodium lauryl sulfate.

Aqueous suspensions of the compounds of formula I contain the active material in admixture with excipients suitable for the manufacture of aqueous suspensions. Such excipients include suspending agents, such as sodium carboxymethylcellulose, croscarmellose, povidone, methylcellulose, hydroxypropylmethylcellulose, sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia; and dispersing or wetting agents such as naturally occurring phosphatides (e.g., lecithin), condensation products of alkylene oxides with fatty acids (e.g., polyoxyethylene stearate), condensation products of ethylene oxide with long chain aliphatic alcohols (e.g., heptadecaethyleneoxycetanol), condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides (e.g., polyoxyethylene sorbitan monooleate). The aqueous suspensions may also contain one or more preservatives, such as ethyl or n-propyl p-hydroxybenzoate, one or more coloring agents, one or more flavoring agents, and one or more sweetening agents, such as sucrose or saccharin.

Pharmaceutical compositions of the compounds of formula I may be in the form of sterile injectable aqueous or oleaginous suspensions, such as sterile injectable aqueous or oleaginous suspensions. This suspension may be formulated according to the methods known in the art using those suitable dispersing or wetting agents and suspending agents which have been mentioned above. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, such as a solution in 1, 3-butanediol or as a lyophilized powder. Acceptable vehicles and solvents that may be used include water, ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any bland fixed oil may be employed including synthetic mono-or diglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables.

For example, a time release formulation intended for oral administration to humans may contain from about 1 to 1000mg of the active compound and a suitable and appropriate amount of carrier material, which may comprise from about 5 to about 95% (weight: weight) of the total composition.

Formulations suitable for parenteral administration include aqueous and non-aqueous sterile injection solutions which may contain antioxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may contain suspending agents and thickening agents.

Formulations suitable for topical administration to the eye also include eye drops wherein the active ingredient is dissolved or suspended in a suitable carrier, especially an aqueous solvent for the active ingredient. The active ingredient is preferably present in the above formulations at a concentration of about 0.5 to 20% w/w, for example about 0.5 to 10% w/w, for example about 1.5% w/w.

Formulations suitable for topical administration in the mouth include lozenges comprising the active ingredient in a flavoured base (usually sucrose and acacia or tragacanth); lozenges comprising the active ingredient in an inert base (such as gelatin and glycerin or sucrose and acacia); and mouthwashes comprising the active ingredient in a suitable liquid carrier.

Formulations suitable for rectal administration may be presented as a suppository with a suitable base comprising, for example, cocoa butter or a salicylate.

Formulations suitable for intrapulmonary or nasal administration have, for example, a particle size of 0.1 to 500 microns (including between 0.1 and 500 microns and in increments such as particle sizes of 0.5, 1, 30, 35 microns, etc.), which are administered as follows: rapid inhalation is through the nasal passages or inhalation is through the mouth to reach the alveolar sacs. Suitable formulations include aqueous or oily solutions of the active ingredient. Formulations suitable for aerosol or dry powder administration may be prepared according to conventional methods and may be delivered with other therapeutic agents, such as compounds heretofore used to treat or prevent the conditions described below.

Formulations suitable for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or spray formulations containing in addition to the active ingredient such carriers as are known in the art to be appropriate.

The formulations may be packaged in unit-dose or multi-dose containers, for example sealed ampoules or vials, and may be stored in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example water, for injections, immediately prior to use. Extemporaneous injection solutions and suspensions are prepared from sterile powders, granules and tablets of the kind described above. Preferred unit dosage formulations are those containing the active ingredient in a daily dose or unit daily sub-dose, or suitable fraction thereof, as herein described.

The present invention also provides a veterinary composition, which thus comprises at least one active ingredient as described above and a veterinary carrier. Veterinary carriers are substances which can be used for the purpose of administering the composition and can be solid, liquid or gaseous substances which are inert or acceptable in the veterinary field and are compatible with the active ingredient. These veterinary compositions may be administered parenterally, orally or by any other desired route.

Combination therapy

The compounds of formula I may be used alone or in combination with other therapeutic agents for the treatment of diseases or disorders described herein, such as inflammatory or hyperproliferative disorders (e.g., cancer). In some embodiments, the compound of formula I is combined in a pharmaceutical combination formulation or administration regimen as a combination therapy with an additional second therapeutic compound that has anti-inflammatory or anti-hyperproliferative properties or that can be used to treat inflammation, immune response disorders, or hyperproliferative disorders (e.g., cancer). The additional therapeutic agent can be a Bcl-2 inhibitor, a JAK inhibitor, an anti-inflammatory agent, an immunomodulator, a chemotherapeutic agent, an apoptosis enhancer, a neurotrophic factor, a cardiovascular disease therapeutic agent, a liver disease therapeutic agent, an antiviral agent, a blood disease therapeutic agent, a diabetes therapeutic agent, and an immunodeficiency disorder therapeutic agent. The second therapeutic agent may be an NSAID anti-inflammatory agent. The second therapeutic agent can be a chemotherapeutic agent. The second compound of the pharmaceutical combination formulation or administration regimen preferably has complementary activities to the compound of formula I such that they do not adversely affect each other. The above compounds are suitably present in combination in amounts effective for the intended purpose. In one embodiment, the compositions of the present application comprise a compound of formula I, or a stereoisomer, tautomer, solvate, metabolite, or pharmaceutically acceptable salt or prodrug thereof, in combination with a therapeutic agent, such as an NSAID.

The combination therapy may be administered on a simultaneous or sequential schedule. When administered first and second, the combination may be administered in two or more administrations. Combined administration includes co-administration and sequential administration in any order using separate formulations or a single pharmaceutical formulation, wherein it is preferred that there is a period of time during which both (or all) active agents exert their biological activities simultaneously.

Suitable dosages for any of the above co-administered drugs are those currently used and may be reduced due to the combined effect (synergy) of the newly identified drug and other therapeutic agent or treatment.

Combination therapy may provide a "synergistic effect" and prove to be "synergistic", i.e. the effect achieved when the active ingredients are used together is greater than the sum of the effects achieved with the compounds separately. When the active ingredients are: (1) co-formulated in a combined unit dose formulation and administered or delivered simultaneously; (2) delivered alternately or in parallel in separate formulations; or (3) when administered by some other regimen, a synergistic effect may be achieved. When delivered in alternation therapy, synergy can be achieved when the compounds are administered or delivered sequentially, e.g., by separate injections in different syringes, separate pills or capsules, or separate infusions. Typically, during alternation therapy, an effective dose of each active ingredient is administered sequentially, i.e., one after the other, while in combination therapy, an effective dose of two or more active ingredients are administered together.

In a specific embodiment of therapy, a compound of formula I, or a stereoisomer, tautomer, solvate, metabolite, or pharmaceutically acceptable salt or prodrug thereof, may be combined with other therapeutic agents, hormonal agents, or antibody agents, such as those described herein, as well as with surgical therapy and radiation therapy. The combination therapies of the present invention thus comprise administering at least one compound of formula I, or a stereoisomer, tautomer, solvate, metabolite, or pharmaceutically acceptable salt or prodrug thereof, and methods of treatment using at least one other cancer. The amounts of the compound of formula I and the other pharmaceutically active therapeutic agent and the associated timing of administration will be selected so as to achieve the desired combined therapeutic effect.

Additional therapeutic agents for use in combination with the compound of formula I include 5-FU, docetaxel, eribulin, gemcitabine, cobicistinib, iptasertib, paclitaxel, tamoxifen, fulvestrant, GDC-0810, dexamethasone, palbociclib, bevacizumab, pertuzumab, trastuzumab-maytansine conjugate, trastuzumab, and letrozole.

Metabolites of compounds of formula I

In vivo metabolites of formula I as described herein also fall within the scope of the present invention. Such products may result, for example, from oxidation, reduction, hydrolysis, amidation, deamidation, esterification, deesterification, enzymatic cleavage, etc. of the administered compound. Accordingly, the present invention includes metabolites of the compounds of formula I, including compounds produced by a method comprising contacting a compound of the present invention with a mammal for a period of time sufficient to produce a metabolite thereof.

Metabolites are typically identified as follows: preparation of the Compounds of the invention radiolabeled (e.g.¹⁴C or³H) Isotopes, which are administered parenterally to animals such as rats, mice, guinea pigs, monkeys, or to humans in detectable doses (e.g., greater than about 0.5mg/kg), allowed a time sufficient for metabolism to occur (typically about 30 seconds to 30 hours) and their conversion products isolated from urine, blood, or other biological samples. These products are easy to isolate because they are labelled (others can be separated by the use of energy)Antibodies that bind to epitopes of antigens that survive the metabolism) metabolite structure is determined in a conventional manner, e.g., by MS, L C/MS or NMR analysis.

Article of manufacture

Another embodiment of the invention provides an article of manufacture or "kit" containing materials useful for the treatment of the above-mentioned diseases and conditions. In one embodiment, the kit comprises a container containing a compound of formula I or a stereoisomer, tautomer, solvate, metabolite, or pharmaceutically acceptable salt or prodrug thereof. The kit may further comprise a label or package insert on or associated with the container. The term "package insert" is used to refer to instructions typically contained in commercial packages of therapeutic products containing information regarding the indications, usage, dosage, administration, contraindications and/or precautions involved in using the above-described therapeutic products. Suitable containers include, for example, bottles, vials, syringes, blister packs, and the like. The container may be formed from a variety of materials such as glass or plastic. The container may contain a compound of formula I or a formulation thereof effective to treat the condition and may have a sterile interface (e.g., the container may be an intravenous solution bag or a vial having a stopper pierceable by a hypodermic injection needle). At least one active agent in the composition is a compound of formula I. The label or package insert indicates that the composition is used to treat the selected condition, e.g., cancer. In addition, the label or package insert may indicate that the patient to be treated is a patient suffering from a condition such as a hyperproliferative condition, neurodegeneration, cardiac hypertrophy, pain, migraine or a neurotrauma disease or event. In one embodiment, the label or package insert indicates that compositions comprising a compound of formula I are useful for treating conditions resulting from abnormal cell growth. The label or package insert may also indicate that the composition can be used to treat other conditions. Alternatively or additionally, the article of manufacture may further comprise a second container comprising a pharmaceutically acceptable buffer, such as bacteriostatic water for injection (BWFI), phosphate buffered saline, ringer's solution, and dextrose solution. It may also contain other substances desirable from a commercial and user standpoint, including other buffers, diluents, filters, needles and syringes.

The kit may also comprise instructions for administering the compound of formula I and the second pharmaceutical formulation (if present). For example, if the kit comprises a first composition comprising a compound of formula I and a second pharmaceutical formulation, the kit may further comprise instructions for administering the first and second pharmaceutical compositions simultaneously, sequentially or separately to a patient in need thereof.

In another embodiment, the kit is suitable for delivering a solid oral form of a compound of formula I, such as a tablet or capsule. The kit preferably comprises a plurality of unit doses. The kit may comprise a card having the dosages arranged in the order of their intended use. An example of such a kit is a "blister pack". Blister packs are known in the packaging industry and are widely used for packaging pharmaceutical unit dose forms. Memory aids, for example in the form of numbers, letters or other indicia or with calendar instructions indicating those days on which administration may be performed in the treatment schedule, may be provided as desired.

According to one embodiment, a kit may comprise (a) a first container having a compound of formula I contained therein; and optionally (b) a second container having a second pharmaceutical formulation contained therein, wherein the second pharmaceutical formulation comprises a second compound having anti-hyperproliferative activity. Alternatively or additionally, the kit may further comprise a third container comprising a pharmaceutically acceptable buffer, such as bacteriostatic water for injection (BWFI), phosphate buffered saline, ringer's solution, and dextrose solution. It may also contain other substances desirable from a commercial and user standpoint, including other buffers, diluents, filters, needles and syringes.

In some other embodiments where the kit comprises a composition of formula I and a second therapeutic agent, the kit may comprise containers for holding separate compositions, such as separate bottles or separate foil packages, although separate compositions may also be contained in a single, undivided container. Typically, the kit comprises instructions for administering the separate components. The kit form is particularly advantageous when the separate components are preferably administered in different dosage forms (e.g. oral and parenteral) or at different dosage intervals or when the attending physician requires titration of the individual components combined.

Preparation of Compounds of formula I

The compounds of formula I can be synthesized by synthetic routes including methods analogous to those known in the chemical arts and particularly in view of the specification of the present application and those for other heterocycles, see Comprehensive Heterocyclic Chemistry II, Katritzky and Rees editions, Elsevier,1997, e.g., Vol.3; L IEbigs Annalen der Chemistry, (9):1910-16, (1985); Helvetica Chimica Acta,41:1052-60, (1958); Arzneimittel-Forschung,40(12):1328-31, (1990), each of which is expressly incorporated as reference raw materials generally available from commercial sources such as Aldrich Chemicals (Milwauk, Wis) or readily prepared using methods known to those skilled in the art (e.g., by the methods in L. eseser and Maresery, supplement 2006, Beijing, German. 9, supplement, Beijing, German-9), and the methods for other heterocycles) (Bessen. 23. supplement, Bessen. 23).

Synthetic chemical transformations and protecting group methodologies (protection and deprotection) and necessary Reagents and intermediates useful in the Synthesis of compounds of formula I are known in the art and are described, for example, in R. L arc, comparative Organic transformations, VCH Publishers (1989), T.W.Greene and P.G.M.Wuts, protective groups in Organic Synthesis, 3 rd edition, John Wiley and Sons (1999), and L. Paquette editors, Encyclopedia of Reagents for Organic Synthesis, John Wiley and Sons (1995) and subsequent versions thereof.

The examples provide exemplary methods of preparing compounds of formula I. One skilled in the art will recognize that other synthetic routes may be used to synthesize the compounds of formula I. Although specific starting materials and reagents are described and discussed in the schemes and examples, other starting materials and reagents can be readily substituted to provide a variety of derivatives and/or reaction conditions. In addition, the various exemplary compounds prepared by the methods described can be further modified based on the present disclosure using conventional chemical methods known to those skilled in the art.

When preparing compounds of formula I, it may be desirable to protect remote functional groups (e.g., primary or secondary amines) of the intermediates. The need for such protection will vary with the nature of the remote functional group and the conditions of the preparation process. Suitable amino protecting groups include acetyl, trifluoroacetyl, tert-Butoxycarbonyl (BOC), benzyloxycarbonyl (CBz) and 9-fluorenylmethyleneoxycarbonyl (Fmoc). The need for such protection is readily determined by one skilled in the art. For a general description of protecting Groups and their use see t.w. greene, Protective Groups in Organic Synthesis, john wiley & Sons, New York, 1991.

In the process for preparing compounds of the formula I, it may be advantageous to separate the reaction products from one another and/or from the starting materials. The desired product of each step or steps is isolated and/or purified to the desired homogeneity by techniques conventional in the art. Typically, such separation involves heterogeneous extraction, crystallization from a solvent or solvent mixture, distillation, sublimation, or chromatography. Chromatography may involve a variety of methods including, for example: reverse phase and normal phase chromatography; size exclusion chromatography; ion exchange chromatography; high, medium and low pressure liquid chromatography and apparatus; small-scale analytical chromatography; simulated Moving Bed (SMB) and preparative thin or thick layer chromatography, and small scale thin and flash chromatography.

Another class of separation methods involves treating a mixture with a reagent selected to bind to or otherwise isolate the desired product, unreacted starting material, reaction by-products, etc. which includes adsorbents or absorbents such as activated carbon, molecular sieves, ion exchange media, etc. alternatively, the reagent can be an acid (in the case of basic materials), a base (in the case of acidic materials), a binding agent such as an antibody, a binding protein, a selective chelating agent such as a crown ether, a liquid/liquid ion extraction reagent (L IX), etc. selection of an appropriate separation method depends on the nature of the material involved, such as boiling point and molecular weight (in distillation and sublimation), the presence or absence of polar functional groups (in chromatography), the stability of the material in acidic and basic media (in heterogeneous extraction), etc.

Enantiomers can be separated by conversion of the enantiomeric mixture to a diastereomeric mixture by reaction with a compound having suitable optical activity (e.g., a chiral auxiliary such as a chiral alcohol or Mosher's acid chloride), separation of the diastereomers, and subsequent conversion (e.g., hydrolysis) of the individual diastereomers to the corresponding pure enantiomers.

Single stereoisomers, such as enantiomers, which are substantially free of their stereoisomers, can be obtained by resolution of a racemic mixture using, for example, optically active resolving agents to form diastereomers (Eliel, E. and Wilen, S. "Stereochemistry of Organic Compounds", John Wiley & Sons, Inc., New York, 1994; L ochmuller, C. H., (1975) J.Chromatogr.,113(3): Fr.302.) the racemic mixture of chiral Compounds of the present application can be separated by any suitable method, including (1) formation of ionic diastereomeric salts with chiral Compounds and separation by fractional crystallization or other Methods, (2) formation of diastereomeric Compounds with chiral derivatizing agents, separation of diastereomers and conversion to pure stereoisomers, and (3) direct separation of substantially pure or enriched stereoisomers under chiral conditions, see "metals, Inc., abstracter, Inc., and abstracts, Inc.

In process (1), diastereomeric salts can be formed by reacting enantiomerically pure chiral bases such as brucine (brucine), quinine, ephedrine, strychnine (strychnine), α -methyl- β -phenylethylamine (amphetamine), etc., with asymmetric compounds bearing acidic functional groups such as carboxylic and sulfonic acids.

Alternatively, by method (2), the substrate to be resolved is reacted with one enantiomer of a chiral compound to form a diastereomer pair (E.and Wilen, S. "Stereochemistry of Organic Compounds", John Wiley&Sons, inc.,1994, p.322.) by reacting an asymmetric compound with an enantiomerically pure derivatizing reagent such as a menthyl derivative to form a diastereomeric compound, which is then separated and hydrolyzed to yield the pure or enriched enantiomer the method of determining optical purity involves preparing a chiral ester of the racemic mixture, such as a menthyl ester, for example, (-) menthyl chloroformate, or Mosher ester, α -methoxy- α - (trifluoromethyl) phenyl acetate (Jacob iii.j.org.chem., (1982)47:4165) in the presence of a base and analyzing the presence of the two atropisomeric enantiomers or diastereomers¹H NMR spectroscopy stable diastereomers of atropisomeric compounds can be separated by normal or reverse phase chromatography followed by separation of the atropisomeric naphthyl-isoquinolines (WO 96/15111) by method (3) racemic mixtures of the two enantiomers can be separated by chromatography using Chiral stationary phases ("Chiral L idchromatograph" (1989) w.j. L ough, ed., Chapman and Hall, New York; Okamoto, j.chromanogr., (1990) 513:375-378) using Chiral stationary phases enriched or purified enantiomers can be distinguished by methods for distinguishing other Chiral molecules with asymmetric carbon atoms, such as optical rotation and circular dichroism.

The compounds of the present invention were prepared as illustrated in general schemes 1-3.

Scheme 1.

a)MgCl₂Heating triethylamine, paraformaldehyde and acetonitrile; b) heating glyoxal and ammonium hydroxide; c) cesium carbonate, 1, 2-dibromoethane, DMF, heating; d) heating N-iodosuccinimide and DMF; e) EtMgBr, THF, -20 ℃ and ii aqueous ammonium chloride solution

As shown in scheme 1, 4-bromo-2-hydroxybenzaldehyde 2 can be obtained by formylation of commercially available 3-bromophenol. Heating with glyoxal 2 gives 3. The formation of the oxaza can be achieved by heating 3 with 1, 2-dibromoethane

And (4) a ring. Bis-iodination can be induced by reaction with N-iodosuccinimide, and the 3-iodo group is selectively removed by treatment with ethyl magnesium bromide at reduced temperature to give 6.

Scheme 2.

f) 4-substituted oxazolidin-2-ones, Cu (OAc)₂trans-N, N' -dimethylcyclohexane-1, 2-diamine, potassium carbonate, dioxane, heating, g) L-proline, CuI, K₃PO₄Heating DMSO; h) ammonium chloride, triethylamine, HATU (1- [ bis (dimethylamino) methylene)]-1H-1,2, 3-triazolo [4,5-b]Pyridinium 3-oxide hexafluorophosphate)

As shown in scheme 2, copper catalysis can be used to couple 6 to the appropriately substituted oxazolidin-2-one to give 7. Brominated intermediate 7 can be coupled to an appropriately substituted amino acid under copper catalysis followed by HATU-mediated amide coupling with ammonium chloride to afford compound 8.

Scheme 3.

i)4,4,5, 5-IVMethyl-2- (tetramethyl-1, 3, 2-dioxaborolan-2-yl) -1,3, 2-dioxaborolan, KOAc, Pd (dppf) Cl₂Heating with dioxane; j) h₂O₂Water, 0 ℃; k) methyl (2R) -2-hydroxypropionate, triphenylphosphine, DEAD, dioxane; l) N-iodosuccinimide, DMF, heating; m) EtMgBr, THF, -40 ℃; n) 4-substituted oxazolidin-2-one, CuI, trans-N, N-dimethyl-1, 2-cyclohexanediamine, potassium carbonate, dioxane, heating; o) ammonia, methanol.

As shown in scheme 3,4 can be converted to boronic acid pinacol ester 9 by palladium catalyzed 4,4,5, 5-tetramethyl-2- (tetramethyl-1, 3, 2-dioxaborolan-2-yl) -1,3, 2-dioxaborane. Oxidation with hydrogen peroxide gives phenol 10. Mitsunobu reaction of 10 with methyl (2R) -2-hydroxypropionate gave 11. Diiodization with N-iodosuccinimide followed by selective removal of the 3-iodo group with ethyl magnesium bromide gives monoiodo 12. Intermediate 12 can be coupled with an appropriately substituted oxazolidin-2-one using copper catalysis to afford oxazolidinone methyl ester 13. The methyl ester of 13 can be converted to the primary amide 14 by heating with ammonia and methanol.

Examples

Abbreviations

DMSO dimethyl sulfoxide

ESI electrospray ionization

HP L C high pressure liquid chromatography

L CMS liquid chromatography mass spectrometry

min for

N equivalent concentration

NMR nuclear magnetic resonance

R_TRetention time

L CMS method A experiments were performed on a Waters Micromass ZQ2000 quadrupole mass spectrometer connected to a Waters Acquity UP L C system with PDA UV detector the spectrometer has an electrospray source operating in positive and negative ion mode the system uses an Acquity BEH C181.7 um 100x 2.1mm column maintained at 40 ℃ or an Acquity BEH Shield RP 181.7 μm 100x 2.1mm column maintained at 40 ℃ and 0.4m L/min flow rate the initial solvent system was 95% water containing 0.1% formic acid (solvent A) and 5% acetonitrile containing 0.1% formic acid (solvent B) for the first 0.4 min, then a gradient to 5% solvent A and 95% solvent B for the next 5.6 min. hold for 0.8 min, then return to 95% solvent A and 5% solvent B in the next 0.2 min. the total run time is 8 min.

L CMS method B experiments were performed on Agilent1100HP L C coupled to an Agilent MSD mass spectrometer using ESI as the ionization source L C separation using a Phenomenex XB-C18,1.7mm, 50 × 2.1.1 mm column with a flow rate of 0.4m L/min, solvent A was water containing 0.1% formic acid, solvent B was acetonitrile containing 0.1% formic acid gradient consisted of 2-98% solvent B over 7 minutes, after equilibration 1.5 minutes 97% B was held for 1.5 minutes L C column temperature was 40 deg.C UV absorbance was collected at 220nm and 254 nm and a full scan of mass spectrum was applied to all experiments.

Example 101(S) -2- ((2- ((R) -4-isopropyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propanamide 101

Step 1: 4-bromo-2-hydroxybenzaldehydes

A 20L 4 neck round bottom flask was purged with nitrogen inert atmosphere and maintained, to which was placed 3-bromophenol (1300g,7.51mol), magnesium dichloride (1078g,11.3mol), triethylamine (3034g,30.0mol) and acetonitrile (7.8L) the mixture was stirred at 40 ℃ for 30 minutes, at 80 ℃ paraformaldehyde (676g,22.6mol) was added to the mixture, the resulting solution was stirred at 76 ℃ for 6 hours the reaction was repeated 5 times, the combined reaction mixture was quenched by the addition of 12L aqueous hydrogen chloride (4N), the pH of the solution was adjusted to 5 with concentrated aqueous hydrogen chloride (12N), the resulting solution was extracted with 1x 20L ethyl acetate, the organic extract was evaporated in vacuo, the residue was purified by silica gel flash chromatography (elution: 15% ethyl acetate/petroleum ether) to give the crude product which was washed with 2.4L methyl tert-butyl ether: hexane (1:4) and the resulting solid was collected by filtration to give 7.0kg of the title compound as a yellow solid.

Step 2: 5-bromo-2- (1H-imidazol-2-yl) phenol

To a 20L 4 neck round bottom flask was placed a solution of 4-bromo-2-hydroxybenzaldehyde (700g,3.50mol) in methanol (7.0L) and acetaldehyde (40%) (2540g,17.5mol) followed by dropwise addition of aqueous ammonia (25-28%, 3500g) over 4 hours with stirring and maintaining the temperature below 40 ℃ the resulting solution was stirred at 30-35 ℃ for 15 hours the reaction was repeated 9 times the combined 9 reaction mixtures were evaporated in vacuo, maintaining the temperature below 45 ℃ the residue was diluted with 100L ethyl acetate, stirring was carried out for 30 minutes the solid was filtered off, the resulting solution was diluted with water, the aqueous phase was extracted with 35L ethyl acetate, the organic extracts were evaporated in vacuo and the residue was purified by silica gel flash chromatography (solvent gradient: 5-75% ethyl acetate/petroleum ether) to give 2.4kg (29%) of the title compound as a yellow solid.

And step 3: 9-bromo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1，4]Oxazazem

To a 20L 4 neck round bottom flask was placed a solution of 5-bromo-2- (1H-imidazol-2-yl) phenol (1.4kg,5.86mol) in N, N-dimethylformamide (14L) and cesium carbonate (7.2kg,22.1 mol). the mixture was stirred for 20 minutes, 1, 2-dibromoethane (4.1kg,21.8mol) was added to the reaction mixture and the resulting solution was stirred at 85-90 deg.C for 4-12 hours, cooled to 15 deg.C, and filtered, the cake was washed with 3.0L ethyl acetate, the filtrate was diluted with 14L ethyl acetate, the combined organic extracts were washed with brine (4X 14L)Washing, drying over anhydrous sodium sulfate, filtering and evaporating in vacuo to give 1.1kg (71%) of the title compound as a pale yellow solid L CMS (ESI) [ M + H ]]⁺＝265；¹H NMR(400MHz,DMSO-d₆)8.32 (d,J＝8.4,1H),7.35-7.24(m,3H),7.06(s,1H),4.47-4.42(m,4H)。

And 4, step 4: 9-bromo-2, 3-diiodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

To a 20L 4 neck round bottom flask was placed 9-bromo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

(2.5kg,9.43mol) and N, N-dimethylformamide (12.5L) were then added in portions with stirring N-iodosuccinimide (6.0kg,26.7 mol.) the resulting solution was stirred at 60 ℃ for 12 hours, cooled to 15 ℃ with a water/ice bath, diluted with 12.5L water/ice and filtered the filtered solid was recrystallized from petroleum ether to give 4.0kg (82%) of the title compound as a yellow solid.

And 5: 9-bromo-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

A20L 4-neck round-bottom flask was purged and maintained under an inert atmosphere of nitrogen, and 9-bromo-2, 3-diiodo-5, 6-dihydrobenzo [ f ] was placed therein]Imidazo [1,2-d ] s][1,4]Oxazazem

(800g,1.55mol) and tetrahydrofuran (2.4L), followed by 3.5 hours at-20 ℃ with stirringEthyl magnesium bromide (1N in ether, 1.7L) was added dropwise the reaction mixture was stirred using an ice/salt bath for 3 hours, the temperature was maintained at-15 ℃. the resulting mixture was quenched by the addition of 3.0L saturated aqueous ammonium chloride and extracted with ethyl acetate (2X 8.0L.) the combined organic extracts were washed with brine (2X 10L), dried over anhydrous sodium sulfate, filtered and evaporated in vacuo, the crude residue was triturated with 8.0L ethyl acetate petroleum ether (1:5), filtered and washed with petroleum ether to give 501g (83%) of the title compound as a brown solid, [ L CMS (ESI): M + H: [ M + H + E ] M + H]⁺＝391；¹HNMR(400MHz,DMSO-d₆)8.22(d,J＝8.7,1H),7.55(s, 1H),7.30-7.25(m,2H),4.45-4.41(m,4H)。

Step 6: (R) -3- (9-bromo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4-isopropyloxazolidin-2-one and (R) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4-isopropyloxazolidin-2-one

Reacting 9-bromo-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

A mixture of (500mg,1.28mmol), (R) -4-isopropyloxazolidin-2-one (231mg,1.79mmol), cuprous iodide (48.7mg, 0.256mmol), trans-N, N' -dimethyl-1, 2-cyclohexanediamine (91 μ L, 0.575mmol) and potassium carbonate (246mg,1.79mmol) was suspended in dioxane (1.25m L) and the reaction mixture was degassed with argon under ultrasonic conditions the resulting mixture was heated at 100 ℃ for 24 h, the reaction was stopped and allowed to cool to room temperature the resulting residue was purified by silica gel flash chromatography (solvent gradient: 0-100% ethyl acetate/cyclohexane) to give 0.965g (about 42%) of the title compound (9-Br: 9)About 1:2 mixture of I products) L CMS (ESI) [ M + H ]]⁺＝391/393/439。

And 7: (R) -3- (9-hydroxy-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4-isopropyloxazolidin-2-one

Reacting (R) -3- (9-bromo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4-isopropyloxazolidin-2-one and (R) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

A mixture of-2-yl) -4-isopropyloxazolidin-2-one (440mg,1.0mmol), cuprous iodide (19.0mg,0.1mmol), (L) -lactic acid (318 μ L, 3.0mmol) and cesium carbonate (1.95g,6.0mmol) is suspended in dimethyl sulfoxide (2.0M L)/water (2.0M L) and the reaction mixture is degassed with argon under ultrasonic conditions.the resulting mixture is heated at 120 ℃ for 24 hours.the reaction is stopped and allowed to cool to room temperature the reaction mixture is neutralized by the addition of 1N hydrochloric acid (6.0M L). the reaction mixture is partitioned between ethyl acetate and brine, the combined organic extracts are dried over magnesium sulfate, filtered and evaporated in vacuo.the crude residue is purified by silica gel flash chromatography (solvent gradient: 0-100% ethyl acetate/cyclohexane) to give 334mg (51%) of the title compound. L CMS (ESI) [ M + H ] ((ESI) ] [ M + H ])]⁺＝330。

And 8: (S) -2- ((2- ((R) -4-isopropyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid methyl ester

Reacting (R) -3- (9-hydroxy-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4-isopropyloxazolidin-2-one (334mg,1.01mmol) dissolved in THF (5.0M L), addition of methyl (2R) -2-hydroxypropionate (145 μ L, 1.52mmol) and triphenylphosphine (399mg,1.52mmol) and stirring of the reaction mixture at room temperature, addition of diisopropyl azodicarboxylate (299 μ L, 1.52mmol) and stirring of the reaction mixture at room temperature for 1.5H, evaporation of the reaction mixture in vacuo and purification of the crude residue by silica gel flash chromatography (solvent gradient: 0-100% ethyl acetate/cyclohexane) to give 421mg (quantitative) of the title compound L cms (esi): M + H]⁺＝416。

And step 9: (S) -2- ((2- ((R) -4-isopropyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid

Mixing (S) -2- ((2- ((R) -4-isopropyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) methyl propionate (421mg,1.00mmol) and lithium hydroxide monohydrate (128mg,3.00mmol) were dissolved in THF/water (4.0M L/2.0M L) and the reaction mixture was set to room temperature with stirring the reaction mixture at room temperature for 1 hour, then neutralized with 1N hydrochloric acid (3.0M L) and vacuum azeotroped with toluene the resulting residue was purified by silica gel flash chromatography (solvent gradient: 0-40% 2N ammonia/methanol/ethyl acetate) to give 260mg (64%) of the title compound L CMS (ESI) [ M + H ] M + H]⁺＝402。

Step 10: (S) -2- ((2- ((R) -4-isopropyl-2-oxooxa-2)Oxazolidin-3-yl) -5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propanamide

(S) -2- ((2- ((R) -4-isopropyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid (260mg,0.647mmol), ammonium chloride (69.3mg, 1.29mmol) and 1- [ bis (dimethylamino) methylene]-1H-1,2, 3-triazolo [4,5-b]Pyridinium 3-oxide hexafluorophosphate (369mg,0.97) was dissolved in N, N-dimethylformamide (5.0m L), N-diisopropylethylamine (332. mu. L, 1.94mmol) was added and the reaction mixture was stirred at room temperature for 1 hour, the resulting mixture was evaporated in vacuo and the residue was purified by silica gel flash chromatography (solvent gradient: 0-10% methanol/ethyl acetate), the product was further purified by reverse phase C18 column (solvent gradient: 0-95% acetonitrile/water, 0.1% ammonium hydroxide buffer) and lyophilized to give 128mg (43%) of 101. L CMS ESI: R_T(min)＝ 3.38,[M+H]⁺401, method a;¹H NMR(400MHz,DMSO-d6)8.12(d,J＝ 8.8Hz,1H),7.50(s,1H),7.23(s,2H),6.68(dd,J＝8.8,2.6Hz,1H),6.46(d,J＝ 2.6Hz,1H),4.60(q,J＝6.6Hz,1H),4.49-4.45(m,1H),4.41-4.33(m,5H), 4.30-4.27(dd,J＝8.9,3.9Hz,1H),2.49-2.47(m,1H),1.40(d,J＝6.6Hz,3H), 0.87(d,J＝7.0Hz,3H),0.74(d,J＝7.0Hz,3H)。

example 102(S) -1- (2- ((R) -4-methyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) pyrrolidine-2-carboxamide 102

Step 1: (R) -4-methyloxazolidin-2-one

At 0 deg.C, to a compound of D-alaninol (8.65g,0.12mmol) in toluene and aqueous KOH (124m L, 12.5% in water, 0.28mmol) was kept at room temperature<Phosgene (72.7m L, 20% in toluene, 0.14mmol) was added at a rate of 5 deg.C, the reaction mixture was stirred at 0 deg.C for a further 40 minutes and then evaporated to dryness, the crude residue was extracted with technical methylated spirit, the slurry was filtered and the filtrate evaporated in vacuo and the residue purified by flash chromatography on silica gel (solvent gradient: 40-100% ethyl acetate/cyclohexane) to give 10.4g (90%) of the title compound as a white solid.¹H NMR(400MHz,CDCl₃)6.00 (br s,1H),4.50(t,J＝6.5Hz,1H),4.07-3.97(m,1H),3.95(dd,J＝7.8,6.2Hz, 1H),1.30(d,J＝6.1Hz,3H)。

Step 2: (R) -3- (9-bromo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4-methyloxazolidin-2-one and (R) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4-methyloxazolidin-2-one

Reacting 9-bromo-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

A mixture of (30.0g,76.7mmol), (R) -4-methyloxazolidin-2-one (7.70g,76.7mmol), cuprous iodide (1.61g,8.40 mmol), trans-N, N' -dimethyl-1, 2-cyclohexanediamine (2.7m L, 16.9mmol) and potassium carbonate (14.9 g,107mmol) was suspended in 1, 4-dioxane (200m L) and the reaction mixture was degassed with argon under sonication conditions the resulting mixture was heated at 100 ℃ for 16 h, the reaction mixture was diluted with aqueous ammonia solution (about 16%) and extracted with ethyl acetate. The combined organic extracts were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo. The resulting residue was purified by silica gel flash chromatography (solvent gradient: 0-100% ethyl acetate/cyclohexane) to yield 13.4g (about 42%) of the title compound (about 2:1 mixture of 9-Br:9-I product).¹H NMR(400MHz,CDCl₃)8.28(d,J＝7.6Hz,0.33H), 8.11(d,J＝6.9Hz,0.66H),7.42-7.38(m,1H),7.28-7.24(m,1.33H),7.23- 7.18(m,0.66H),4.77-4.68(m,1H),4.58(t,J＝8.3Hz,1H),4.49-4.39(m,2H), 4.37-4.30(m,2H),4.08(dd,J＝8.4,4.5Hz,1H),1.57-1.50(m,3H)。

And step 3: (R) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4-methyloxazolidin-2-one

Separation of 80mg of (R) -3- (9-bromo-5, 6-dihydrobenzo [ f ] by chiral SFC]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4-methyloxazolidin-2-one and (R) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

A mixture of (E) -2-yl) -4-methyloxazolidin-2-one to yield 35.0mg of the title compound L CMS (ESI) [ M + H ]]⁺＝412.0；¹H NMR(400MHz,DMSO-d6)8.04 (d,J＝8.5Hz,1H),7.47(d,J＝1.8Hz,1H),7.45(d,J＝1.8Hz,1H),7.41(d,J＝ 1.8Hz,1H),7.35(s,1H),4.64-4.54(m,2H),4.47-4.41(m,4H),4.09-4.06(m, 1H),1.41(d,J＝6.0Hz,3H)。

And 4, step 4: (2- ((R) -4-methyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) -L-proline

Reacting (R) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

A mixture of-2-yl) -4-methyloxazolidin-2-one (414mg,1.01mmol), L-proline (579mg,5.03mmol), cuprous iodide (81 mg,0.42mmol) and tripotassium phosphate (128mg,6.03mmol) in dimethyl sulfoxide (4.0m L) was heated at 80 deg.C for 18 hours crude reaction mixture was passed through

SCX-2 column purification followed by further purification by silica gel flash chromatography (solvent gradient: 0-30% 2N ammonia/methanol/dichloromethane) gave 262mg (65%) of the title compound L CMS (ESI) [ M + H ]]⁺＝399。

And 5: (S) -1- (2- ((R) -4-methyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) pyrrolidine-2-carboxamide

To (2- ((R) -4-methyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) -L-proline (262mg,0.66mmol), ammonium chloride (70mg,1.31mmol) and N, N-diisopropylethylamine (0.34m L, 1.97mmol) in N, N-dimethylformamide (3.0m L) 1- [ bis (dimethylamino) methylene ] chloride was added]-1H-1,2, 3-triazolo [4,5-b]Pyridinium 3-oxide hexafluorophosphate (374mg,0.98mmol) and the reaction mixture stirred at room temperature for 6 hours the resulting mixture was evaporated in vacuo, purified by silica gel flash chromatography (solvent gradient: 0-10% methanol/ethyl acetate) and then further purified by reverse phase HP L C and lyophilized to give 110mg (42%) 102 as a white solid L CMS (ESI) R_T(min)＝2.60[M+H]⁺398.0, method a;¹H NMR(400 MHz,DMSO-d₆)8.03(d,J＝8.8Hz,1H),7.37(br s,1H),7.12(s,1H),7.02(br s,1H),6.28(dd,J＝8.9,2.4Hz,1H),5.99(d,J＝2.4Hz,1H),4.58-4.47(m, 2H),4.38-4.25(m,4H),4.05-3.96(m,1H),3.92-3.87(m,1H),3.56-3.47(m, 1H),3.24-3.16(m,1H),2.21-2.10(m,1H),2.00-1.86(m,3H),1.37(d,J＝ 5.9Hz,3H)。

example 103(S) -1- (2- ((S) -2-oxo-4- (trifluoromethyl) oxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) pyrrolidine-2-carboxamide 103

Step 1: (R) -2- ((tert-butyldimethylsilyl) oxy) -1-phenyleth-1-amine

To a solution of (R) -2-amino-2-phenylethanol (50g,0.36mol) in dichloromethane (500m L) was added 4- (dimethylamino) pyridine (9.0g,73.6mmol) and triethylamine (102m L, 0.73 mol). the reaction mixture was cooled in an ice bath and a solution of tert-butyldimethylchlorosilane (54.8g,0.36mol) in dichloromethane (300m L) was slowly added, maintaining the internal temperature<10 ℃. The reaction mixture was allowed to warm to ambient temperature and stirred overnight. The resulting mixture was washed twice with water and then brine, dried over sodium sulfate, filtered and evaporated in vacuo. The crude oil was purified by silica gel flash chromatography (solvent gradient: 5-50% ethyl acetate/cyclohexane) to give 65.0g (71%) of the title compound as a colorless oil.¹H NMR(400 MHz,CDCl₃)7.38-7.23(m,5H),4.07(dd,J＝8.4,3.9Hz,1H),3.72(dd,J＝ 9.7,3.9Hz,1H),3.51(dd,J＝9.5,8.5Hz,1H),1.72(br s,2H),0.90(s,9H),0.02 (s,6H)。

Step 2: (R) -2- ((2- ((tert-butyldimethylsilyl) oxy) -1-phenylethyl) imino) -3,3, 3-trifluoropropionic acid ethyl ester

A compound of (R) -2- ((tert-butyldimethylsilyl) oxy) -1-phenylethane-1-amine (63.5g,253 mmol), ethyl 3,3, 3-trifluoro-2-oxopropanoate (48.5g,285mmol) and pyridinium p-toluenesulfonate (6.76g,36.9mmol) in toluene (2.0L) was heated under Dean-Stark reflux conditions for 16 h, the reaction mixture was cooled to ambient temperature and evaporated in vacuo, ether (3.75L) was added to the resulting residue and the insoluble solids were removed by filtration and the resulting ether solution was used in the next step without purification.

And step 3: (S) -3,3, 3-trifluoro-2- (((R) -2-hydroxy-1-phenylethyl) amino) propan-1-ol

To ethyl (R) -2- ((2- ((tert-butyldimethylsilyl) oxy) -1-phenylethyl) imino) -3,3, 3-trifluoropropionate in diethyl ether (crude from step 2 above, assumed to be 253mmol) was slowly added lithium aluminium hydride (755m L, 1.0N solution in tetrahydrofuran, 755mmol) at 0 deg.C, maintaining the temperature<The reaction mixture was allowed to warm to ambient temperature and stirred for 16 h, quenched by adding dropwise water (31m L), then NaOH (31m L, 15% aq), followed by water (93m L), and held at temperature<10 ℃. The resulting precipitate was removed by filtration through celite and the filtrate was passed through a pad of silica gel. The filtrate was evaporated in vacuo and the resulting residue was purified by silica gel flash chromatography (solvent gradient: 10-80% ethyl acetate/cyclohexane) to give 28g (45%) of the title compound (first eluted isomer) as a white solid.¹H NMR(400 MHz,CDCl₃)7.39-7.30(m,5H),4.15(dd,J＝9.0,4.1Hz,1H),3.75(dd,J＝ 10.7,4.0Hz,1H),3.68-3.54(m,3H),3.14-3.06(m,1H),2.43(br s,1H),2.22 (br s,1H),2.04(br s,1H)。

And 4, step 4: (S) -2-amino-3, 3, 3-trifluoropropan-1-ol hydrochloride

To a solution of (S) -3,3, 3-trifluoro-2- (((R) -2-hydroxy-1-phenylethyl) amino) propan-1-ol (28.0g,112mmol) in ethanol (300m L) were added HCl (118m L, 1.25N in methanol) and HCl (28.4m L, 4N in dioxane), followed by palladium hydroxide on carbon (3.86g,20 wt%) and the reaction mixture was stirred under an atmosphere of hydrogen for 6.5 hours the reaction mixture was filtered and the filtrate evaporated in vacuo the residue triturated with ether to give 17.1g (92%) of the title compound as a white solid [ α% ]]_D-9 ° (c ═ 2.0, ethanol);¹H NMR(400MHz,MeOD-d₄)4.19-4.11(m,1H),3.96(dd,J＝ 12.3,4.3Hz,1H),3.89(dd,J＝12.1,5.9Hz,1H)。

and 5: (S) -4-trifluoromethyl oxazolidin-2-one

To a vigorously stirred mixture of (S) -2-amino-3, 3, 3-trifluoropropan-1-ol hydrochloride (12.6g,76.1mmol) and KOH (12.5% aq,175m L, 0.39mol) in toluene (85m L) was added phosgene (78m L, 20% solution in toluene, 0.157mmol) dropwise at 13 deg.C, maintaining the temperature<The reaction mixture was stirred at 10 ℃ for a further 15 minutes and then evaporated in vacuo, the residual solid was extracted with hot technical methylated ethanol, filtered through a pad of celite and the filtrate evaporated in vacuo, the crude residue was purified by flash chromatography on silica gel (solvent gradient: 30-100% ethyl acetate/cyclohexane) to give 8.87g (75%) of the title compound as a white solid [ α ]]_D+18°(c＝1.0,methanol)；1H NMR(400MHz,CDCl₃)6.01 (br s,1H),4.58(t,J＝9.4Hz,1H),4.51(dd,J＝9.7,3.9Hz,1H),4.35-4.27(m, 1H)。

Step 6: (S) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4- (trifluoromethyl) oxazolidin-2-one and (S) -3- (9-bromo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4- (trifluoromethyl) oxazolidin-2-one

Reacting 9-bromo-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

(24.6g,62.9mmol), (S) -4-trifluoromethyloxazolidin-2-one (8.87g,57.2mmol), cuprous iodide (2.2g,11.4 mmol), trans-N, N' -dimethylcyclohexane-1, 2-diamine (3.6m L, 22.9mmol) and potassium carbonate (15.8 g,114mmol) were suspended in dioxane (200m L) and degassed with argon under ultrasonic conditions the reaction mixture was heated at 100 ℃ for 16 h, the resulting mixture was diluted with aqueous ammonia (about 16%) and extracted with ethyl acetate, the combined organic extracts were washed with brine, dried over magnesium sulfate and evaporated in vacuo, the crude residue was purified by silica gel flash chromatography (solvent gradient: 0-100% ethyl acetate/cyclohexane) to give 8.11g (about 31%) of the title compound (7: 3 mixture of 9-Br product).¹H NMR(400MHz, DMSO-d₆)8.22(d,J＝8.6Hz,0.3H),8.04(d,J＝8.6Hz,0.7H),7.49-7.44(m, 0.7H),7.42-7.41(m,1.7H),7.34-7.31(m,0.3H),7.27-7.26(m,0.3H),5.55- 5.47(m,1H),4.76-4.71(m,1H),4.66-4.63(m,1H),4.50-4.41(m,4H)。

And 7: (2- ((S) -2-oxo-4- (trifluoromethyl) oxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) -L-proline

Reacting (S) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4- (trifluoromethyl) oxazolidin-2-one and (S) -3- (9-bromo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

A mixture of-2-yl) -4- (trifluoromethyl) oxazolidin-2-one (960mg, 7:3 mixture of 9-I:9-Br, ca. 2.19mmol), L-proline (630mg,5.48mmol), cuprous iodide (84mg,0.44mmol) and tripotassium phosphate (1.86g, 8.88mmol) in dimethyl sulfoxide (7.0m L) was heated at 100 ℃ for 18 hours the cooled reaction mixture was added to stirred dichloromethane (140m L) and flash chromatographed on silica gel (solvent gradient: 0-35% 2N NH 3)₃MeOH/dichloromethane) to yield 262mg (65%) of the title compound L CMS (ESI) [ M + H ]]⁺＝453。

And 8: (S) -1- (2- ((S) -2-oxo-4- (trifluoromethyl) oxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) pyrrolidine-2-carboxamide

To ice-cooled (2- ((S) -2-oxo-4- (trifluoromethyl) oxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] an]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) -L-proline (582mg,1.29 mmol), ammonium chloride (138mg,2.57mmol) and N, N-diisopropylethylamine (1.0m L, 5.80mmol) in DMF (5.0m L) was added 1- [ bis (dimethylamino) methylene ] amine in portions]-1H-1,2, 3-triazolo [4,5-b]Pyridinium 3-oxide hexafluorophosphate (1.22g,3.21 mmol). The reaction mixture was stirred at room temperature for 15 minutes and then partitioned between ethyl acetate and dilute aqueous sodium bicarbonate. The aqueous phase was extracted with ethyl acetate and the combined organic extracts were washed successively with water, brine, dried over sodium sulfate and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (solvent gradient: 0-10% methanol/dichloromethane) and further on silica gel (solvent gradient: 0-80% methyl acetate/ethyl acetate)Purification and final recrystallization from acetonitrile gave 245mg (42%) 103 as a pale yellow solid L CMS (ESI)_T(min)＝3.34[M+H]⁺452.2, method a;¹H NMR(400MHz,DMSO-d₆)8.06 (d,J＝8.9Hz,1H),7.40(br s,1H),7.21(s,1H),7.05(brs,1H,),6.32(dd,J＝ 8.9,2.4Hz,1H),6.03(d,J＝2.4Hz,1H),5.50-5.42(m,1H),4.73(t,J＝9.3Hz, 1H),4.62(dd,J＝10.1,2.4Hz,1H),4.40-4.34(m,4H),3.96-3.93(m,1H), 3.58-3.53(m,1H),3.27-3.20(m,1H),2.29-2.15(m,1H),2.03-1.91(m, 3H)。

example 104(S) -2- ((2- ((R) -4-ethyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propanamide 104

Step 1: (R) -4-Ethyl-3- (9-bromo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) oxazolidin-2-one and (R) -4-ethyl-3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) oxazolidin-2-ones

Reacting 9-bromo-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

A mixture of (1.5g,3.84mmol), (R) -4-ethyloxazolidin-2-one (618mg,5.37mmol), cuprous iodide (146mg,0.767 mmol), trans-N, N' -dimethyl-1, 2-cyclohexanediamine (272. mu. L, 1.73mmol) and potassium carbonate (742 mg,5.37mmol) was mixedSuspended in dioxane (4.5M L) and the reaction mixture degassed with argon under sonication the resulting mixture was heated at 100 ℃ for 24H, the reaction was stopped and allowed to cool to room temperature the residue was purified by flash chromatography on silica gel (solvent gradient: 0-100% ethyl acetate/cyclohexane) to give 680mg (42%) of the title compound (about 1:2 mixture of 9-Br:9-I product): L cms (esi): M + H]⁺＝378/380/426。

Step 2: (R) -4-Ethyl-3- (9-hydroxy-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) oxazolidin-2-ones

Reacting (R) -4-ethyl-3- (9-bromo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) oxazolidin-2-one and (R) -4-ethyl-3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) oxazolidin-2-one (680mg,1.6mmol), cuprous iodide (30.5mg,0.16mmol), (L) -lactic acid (421 μ L, 4.80mmol) and cesium carbonate (3.12g,6.01mmol) were suspended in dimethyl sulfoxide (3.0M L)/water (3.0M L) and the reaction mixture was degassed with argon under ultrasonic conditions the resulting mixture was heated at 120 ℃ for 24H stopping the reaction and the reaction mixture was cooled to room temperature, neutralized by addition of 1N hydrochloric acid (6.0M L) and partitioned between ethyl acetate and brine, the combined organic extracts were dried over magnesium sulfate, filtered and evaporated in vacuo, the resulting residue was purified by silica gel flash chromatography (solvent gradient: 0-100% ethyl acetate/cyclohexane) to give 196mg (39%) of the title compound L CMS (ESI): M + H: [ M + H ESI ] (M + H + E)]⁺＝316。

And step 3: (S) -2- ((2- ((R) -4-ethyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid methyl ester

Reacting (R) -4-ethyl-3- (9-hydroxy-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) oxazolidin-2-one (196mg,0.623mmol) dissolved in THF (5.0M L) was added methyl (2R) -2-hydroxypropionate (89. mu. L, 0.934mmol) and triphenylphosphine (245mg,0.934mmol) and the reaction mixture was set to stir at room temperature then diisopropyl azodicarboxylate (183. mu. L, 0.934mmol) was added and the reaction mixture was stirred at room temperature for 1.5H the reaction mixture was evaporated in vacuo and the resulting residue was purified by silica gel flash column chromatography (solvent gradient: 0-100% ethyl acetate/cyclohexane) to give 214mg (86%) of the title compound. L (CMS ESI): M + H ESI [ M + H ESI ])]⁺＝402。

And 4, step 4: (S) -2- ((2- ((R) -4-ethyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propanamide

Reacting (S) -2- ((2- ((R) -4-ethyl-2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-Yl) oxy) methyl propionate (214mg,0.534mmol) was dissolved in 7N ammonia/methanol (20m L, 140 mmol.) the reaction mixture was stirred at room temperature for 3 hours the resulting mixture was evaporated in vacuo, purified by reverse phase C18 chromatography (solvent gradient: 0-95% acetonitrile/water, 0.1% ammonium hydroxide buffer) and lyophilized to give 163mg (79%) of 104. L CMS (ESI): R_T(min)＝3.12,[M+H]⁺387, method a;¹H NMR(400MHz,DMSO-d₆)8.13(d,J＝8.9Hz,1H),7.50 (s,1H),7.21(s,2H),6.68(dd,J＝8.8,2.6Hz,1H),6.46(d,J＝2.6Hz,1H),4.61 (q,J＝6.6Hz,1H),4.52-4.46(m,2H),4.40-4.32(m,4H),4.20-4.15(m,1H),1.85-1.76(m,2H),1.40(d,J＝6.6Hz,3H),0.80(t,J＝7.5Hz,3H)。

example 105(S) -2- ((2- ((S) -2-oxo-4- (trifluoromethyl) oxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propanamide 105

Step 1: 9- (4,4,5, 5-tetramethyl-1, 3, 2-dioxolan-2-yl) -5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

A3L 4-neck round-bottom flask was purged and maintained with a nitrogen inert atmosphere, and dioxane (1.5L), 9-bromo-5, 6-dihydrobenzo [ f ] was placed therein]Imidazo [1,2-d ] s][1,4]Oxazazem

(150g,566mmol), 4,5, 5-tetramethyl-2- (tetramethyl-1, 3, 2-dioxolan-2-yl) -1,3, 2-dioxolan-e (217 g,855mmol), potassium acetate (165g,1.68mol,2.97 eq.) and Pd (dppf) Cl₂(42g,57.4 mmol). The resulting solution was stirred at 90 ℃ overnight. The resulting solid was filtered and the filtrate was evaporated in vacuo. The crude solid obtained is purified by flash chromatography on silica gel (eluent: 25% ethyl acetate/petroleum ether) to yield 154g (87%) of the title compound as a pale yellow oil.

Step 2:5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-alcohols

Into a 5L 4 neck round bottom flask were placed dichloromethane (1.0L) and 9- (4,4,5, 5-tetramethyl-1, 3, 2-dioxolan-2-yl) -5, 6-dihydrobenzo [ f ] n at 0 ℃ with stirring]Imidazo [1,2-d ] s][1,4]Oxazazem

(100g,320mmol) followed by the dropwise addition of 30% aqueous hydrogen peroxide (700m L, 8.82 mol.) the reaction mixture was stirred overnight at room temperature, the resulting mixture was taken up with Na₂SO₃The resulting solution was extracted with ethyl acetate (4 × 5L), the organic extracts combined, dried over anhydrous sodium sulfate, filtered and evaporated in vacuo to give 216g (crude, solvent-containing) of the title compound as a white solid which was used without further purification.

And step 3: (S) -2- ((5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid methyl ester

A5L 4-neck round-bottom flask was purged and maintained under an inert atmosphere of nitrogen, and dioxane (3.0L), 5, 6-dihydrobenzo [ f ] was placed therein]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-ol (136g,673mmol), (2R) -methyl 2-hydroxypropionate (91g,874mmol), triphenylphosphine (265g,1.01 mol). Diethyl azodicarboxylate (176g,1.01mol) was then added over 1 hour at 10 ℃. The resulting solution was stirred at room temperature overnight. The reaction mixture was evaporated in vacuo and the crude product was flash chromatographed on silica gel (elution)Preparation: 50% ethyl acetate/petroleum ether) to yield 170g (88%) of the title compound as a yellow solid.

And 4, step 4: (S) -2- ((2, 3-diiodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid methyl ester

A5L 4-neck round-bottom flask was purged with and maintained under an inert atmosphere of nitrogen, and N, N-dimethylformamide (3.0L), (S) -2- ((5, 6-dihydrobenzo [ f ] was placed therein]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) methyl propionate (335g,1.16mol) and N-iodosuccinimide (262g,1.16 mol.) the reaction mixture is stirred at 50 ℃ for 3 hours then the resulting solution is quenched by the addition of 5L water/ice the solid is collected by filtration, extracted with 5L ethyl acetate and evaporated in vacuo the crude product is purified by flash chromatography on silica gel (eluent: 15% ethyl acetate/petroleum ether) to give 200g (32%) of the title compound as a yellow solid.

And 5: (S) -2- ((2-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid methyl ester

A10L 4-neck round-bottom flask was purged and maintained with a nitrogen inert atmosphere, to which was placed tetrahydrofuran (5L) and (S) -2- ((2, 3-diiodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid methyl ester (296g,548mmol) The reaction mixture was stirred at-40 ℃ for 4 hours, the resulting solution was quenched by addition of 1L saturated aqueous ammonium chloride and extracted with ethyl acetate (2x 3L), the combined organic extracts were dried over anhydrous sodium sulfate, filtered and evaporated in vacuo, the crude product was purified by silica gel flash chromatography (solvent gradient: 25-40% ethyl acetate/petroleum ether) to give 131g (58%) of the title compound, L CMS ESI, [ M + H)]⁺＝415；¹H NMR(400MHz,CDCl₃)8.43(d,J＝4.5Hz,1H),7.00(s,1H),6.67- 6.63(m,1H),6.49(d,J＝1.2Hz,1H),4.81-4.42(m,1H),4.41-4.39(m,2H), 4.39-4.30(m,2H),3.76(s,3H),1.63(d,J＝3.45Hz,3H)。

Step 6: (S) -2- ((2- ((S) -2-oxo-4- (trifluoromethyl) oxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid methyl ester

Reacting (S) -2- ((2-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

A mixture of methyl (1.33g,3.20mmol), (S) -4-trifluoromethyloxazolidin-2-one (500mg,3.20mmol), cuprous iodide (240mg,1.30mmol), trans-N, N' -dimethylcyclohexane-1, 2-diamine (200mg, 1.30mmol) and potassium carbonate (880mg,640mmol) in dioxane (20M L) was degassed with argon under ultrasonic conditions and the reaction mixture was heated at 100 ℃ for 6 hours the resulting mixture was diluted with water and extracted with ethyl acetate the combined organic extracts were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo the crude residue was purified by silica gel flash chromatography (solvent gradient: 0-100% ethyl acetate/cyclohexane) to give 890mg (63%) of the title compound as a white solid L (esi) ("M + H]⁺＝442。

And 7: (S) -2- ((2- ((S) -2-oxo-4- (trifluoromethyl) oxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propanamide

Reacting (S) -2- ((2- ((S) -2-oxo-4- (trifluoromethyl) oxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid methyl ester, (S) -2- ((2- ((S) -4- (trifluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

A solution of-9-yl) oxy) methyl propionate (890mg,2.01mmol) in ammonia solution (10m L, 7N in methanol, 70mmol) was stirred at room temperature for 18 h the reaction mixture was evaporated in vacuo and the resulting solid recrystallized from dichloromethane and dried in vacuo to give 562mg (65%) 105 as a white solid L CMS (ESI)_T(min)＝3.41,[M+H]⁺427, method a;¹H NMR(400MHz,DMSO-d₆)8.19 (d,J＝9.1Hz,1H),7.54(s,1H),7.30(s,1H),7.26(s,1H),6.73(dd,J＝9.0,2.5 Hz,1H),6.51(d,J＝2.6Hz,1H),5.53–5.44(m,1H),4.74(t,J＝9.6Hz,1H), 4.69-4.61(m,2H),4.48-4.38(m,4H),1.43(d,J＝6.6Hz,3H)。

example 106(S) -1- (2- ((S) -4- (difluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) pyrrolidine-2-carboxamide 106

Step 1: (R) -2, 2-dimethyl- [1,3] dioxolane-4-carbaldehyde

Sodium periodate (57.0g,270mmol) was dissolved in hot water (115m L) and silica gel (200g,

220-440 mesh, particle size 35-75 μm) the mixture was stirred vigorously until a free-flowing powder was obtained, it was added to a solution of 1,2:5, 6-di-O- (1-methylethylidene) -D-mannitol (50g,190mmol) in dichloromethane (1.0L) and the reaction mixture was stirred at room temperature for 1 hour₂SO₄The pad was filtered and the solid was washed thoroughly with dichloromethane. The combined organic extracts were evaporated in vacuo to give 37.2g (75%) of the title compound as a colourless oil.¹H NMR(400MHz,CDCl₃) 9.73(d,J＝1.9Hz,1H),4.38(ddd,J＝7.4,4.7,1.9Hz,1H),4.18(dd,J＝8.8,7.4Hz,1H),4.10(dd,J＝8.8,4.7Hz,1H),1.49(s,3H),1.43(s,3H)。

Step 2: (R) -4-difluoromethyl-2, 2-dimethyl- [1,3] dioxolane

To (R) -2, 2-dimethyl- [1,3] cooled in a water bath]Dioxolane-4-carbaldehyde (7.08g,54mmol) was added dropwise to a solution of diethylaminosulfur trifluoride (8.4m L, 62.6 mmol) in dichloromethane (50m L) and the reaction mixture was stirred at room temperature for 3 hours the resulting mixture was added dropwise to a rapidly stirred, ice-cold saturated aqueous sodium bicarbonate solution the mixture was further extracted with dichloromethane the combined organic extracts were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo to give 6.58g (79%) of the crude title compound as an orange oil.¹H NMR(400MHz,CDCl₃)5.69(td,J＝ 55.8,4.9Hz,1H),4.27-4.17(m,1H),4.16-4.03(m,2H),1.46(s,3H),1.38(s,3H)。

And step 3: (R) -3- (tert-butyldimethylsilyloxy) -1, 1-difluoropropan-2-ol

HCl/dioxane (4N,10.8m L, 43.2mmol) was added to (R) -4-difluoromethyl-2, 2-dimethyl [1,3] -dimethyl]Dioxolane (6.58g,43.2mmol) in methanol (40m L) and the reaction mixture stirred at room temperature for 30 minutes the resulting mixture was evaporated in vacuo and azeotroped with acetonitrile the residue was dissolved in N, N-dimethylformamide (10m L) and tert-butyldimethylchlorosilane (6.53g,43.2 mmol), triethylamine (9.0m L, 64.9mmol) and 4- (dimethylamino) pyridine) (catalytic amounts) were added the reaction mixture stirred at room temperature for 1 hour the resulting mixture was washed with water then extracted with dichloromethane the combined organic extracts washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo the crude residue obtained was purified by flash chromatography on silica gel (solvent gradient: 0-30% ethyl acetate/cyclohexane) to give 3.43g (35%) of the title compound as a yellow oil.¹H NMR(400MHz,CDCl₃)5.66(td, J＝56.4,4.6Hz,1H),3.76-3.60(m,2H),2.46(d,J＝6.4Hz,1H),0.81(s,9H), 0.00(s,6H)。

And 4, step 4: ((S) -2-azido-3, 3-difluoropropoxy) -tert-butyldimethylsilane

Triflic anhydride (2.9m L, 17.4mmol) was added dropwise to a solution of (R) -3- (tert-butyldimethylsilyloxy) -1, 1-difluoropropan-2-ol (3.43g,15.1mmol) and pyridine (2.0m L, 24.2 mmol) in dichloromethane (50m L) at-20 ℃ and the reaction mixture was stirred at-20 ℃ for 20 minutes and then at 0 ℃ for 1 hour the resulting mixture was diluted with 0.5N aqueous HCl and extracted with dichloromethane the combined organic extracts were dried over magnesium sulfate and evaporated in vacuo the crude residue was dissolved in N, N-dimethylformamide (10m L), sodium azide (2.96g,45.5mmol) was added and the reaction mixture was stirred at room temperature for 2 hours the resulting mixture was diluted with water and extracted with dichloromethaneExtraction was performed with ethyl acetate. The combined organic extracts were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo to give 4.50g of the crude title compound.¹H NMR(400MHz,CDCl₃)5.74(td,J＝55.4,4.4Hz,1H),3.81- 3.71(m,2H),3.58-3.47(m,1H),0.81(s,9H),0.00(s,6H)。

And 5: (S) -1- (tert-butyldimethylsilyloxymethyl) -2, 2-difluoroethylamine

Palladium hydroxide on carbon (200mg, 20%) was added to a solution of ((R) -2-azido-3, 3-difluoropropoxy) -tert-butyldimethylsilane (4.50g, crude, assumed to be about 15.1mmol) in ethyl acetate (20m L) and methanol (2.0m L) and the reaction mixture was stirred under a hydrogen balloon for 16 h the reaction mixture was filtered, fresh palladium hydroxide on carbon (400mg, 20%) was added and the reaction mixture was stirred under a hydrogen balloon for 16 h the resulting mixture was filtered and the filtrate was evaporated in vacuo to give 3.08g (90%) of the crude title product as a colorless oil.¹H NMR(400MHz,CDCl₃)5.66(td,J＝57.0,4.7 Hz,1H),3.71-3.57(m,2H),3.00-2.89(m,1H),1.42(br s,2H),0.82(s,9H),0.00(s,6H)。

Step 6: (S) -4-difluoromethyl oxazolidin-2-one

HCl/dioxane (4N,5.0m L, 20mmol) was added to a solution of (R) -1- (tert-butyldimethylsilyloxymethyl) -2, 2-difluoroethylamine (Org. L et t., vol.9, No.1,2007,41-44) (2.30g, 10.3mmol) in methanol (5.0m L) and the reaction mixture was stirred at room temperature for 2 hours the mixture was evaporated in vacuo, the resulting oil was triturated with diethyl ether to give a solid which was dried in vacuo, the solid was dissolved at 0 ℃ in a mixture of toluene (20m L) and KOH (2.50g,44.6mmol in 20m L water), phosgene (16.3m L, 20% in toluene) was added dropwise, the cooling bath was removed and the reaction mixture was stirred for 1 hour the mixture was evaporated in vacuo,the resulting residue was extracted with hot industrial methylated ethanol, the solid was collected by filtration, the filtrate was evaporated in vacuo and the resulting residue was purified by flash chromatography on silica gel (solvent gradient: 0-100% ethyl acetate/cyclohexane) to give 830mg (68%) of the title compound as an off-white solid [ α ]]_D＝+10.1(c＝2.37,CHCl₃)。¹H NMR(400MHz,CDCl₃)5.96(br s,1H), 5.78(td,J＝55.3,4.8Hz,1H),4.54(t,J＝9.2Hz,1H),4.42(dd,J＝9.6,4.4Hz, 1H),4.17-4.06(m,1H)。

And 7: (S) -4- (difluoromethyl) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) oxazolidin-2-ones

Reacting 9-bromo-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

(741mg,1.90mmol), (S) -4-difluoromethyl oxazolidin-2-one (260mg,1.90mmol), cuprous iodide (72mg, 0.38mmol), trans-N, N' -dimethylcyclohexane-1, 2-diamine (120. mu. L, 0.76mmol), and potassium carbonate (524mg,3.79mmol) in dioxane (10M L) were degassed with argon under sonication conditions the reaction mixture was heated at 100 ℃ for 16H the resulting mixture was diluted with water and extracted with ethyl acetate the combined organic extracts were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo the crude residue was purified by silica gel flash chromatography (solvent gradient: 0-100% ethyl acetate/cyclohexane) to give 438mg (50%) of the title compound L CMS (ESI): M + H]⁺＝448；¹H NMR(400MHz, CDCl₃)8.03(d,J＝8.7Hz,1H),7.44-7.39(m,2H),7.29(s,1H),6.64(t,J＝ 56.8Hz,1H),4.86(ddd,J＝24.0,9.2,4.0Hz,1H),4.73(dd,J＝9.3,4.0Hz,1H), 4.53(t,J＝9.3Hz,1H),4.46-4.41(m,2H),4.38-4.32(m,2H)。

And 8: (2- ((S) -4- (difluoromethyl) -2-oxoOxazolidin-3-yl) -5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) -L-proline

Reacting (S) -4- (difluoromethyl) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

A mixture of-2-yl) oxazolidin-2-one (218mg,0.49mmol), L-proline (112mg,0.97mmol), cuprous iodide (19mg,0.10mmol) and tripotassium phosphate (207mg,0.98mmol) in dimethyl sulfoxide (2.0m L) was degassed with argon under ultrasonic conditions, the reaction mixture was heated at 100 ℃ for 16 hours, the resulting mixture was diluted with dichloromethane (30m L) and then loaded directly onto a silica gel flash column (10 g) and eluted (solvent gradient: 5-30% 2N NH)₃Methanol/dichloromethane) to yield the crude title compound L CMS (ESI) [ M + H ]]⁺＝435。

And step 9: (S) -1- (2- ((S) -4- (difluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) pyrrolidine-2-carboxamide

1- [ bis (dimethylamino) methylene]-1H-1,2, 3-triazolo [4,5-b]Pyridinium 3-oxide hexafluorophosphate HATU (259mg,0.68mmol) was added portionwise to (S) -1- (2(2- ((S) -4- (difluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f [ -f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) -L-proline (crude, ca. 0.49mmol), ammonium chloride (52mg,0.97mmol) and triethylamine (136. mu. L, 0.97mmol) in N, N-dimethylformamide (2.0m L) and the reaction mixture was stirred at room temperature for 15 minAqueous sodium bicarbonate was diluted and extracted with ethyl acetate. The combined organic extracts were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo. Flash chromatography of the crude residue on silica gel (solvent gradient: 0-10% 2N NH)₃Methanol/dichloromethane) to yield 82mg (39%) 106 as an off-white solid L CMS (ESI)_T(min)＝3.31,[M+H]⁺434, method a;¹H NMR(400MHz,DMSO-d₆)8.05(d,J＝8.9Hz,1H),7.36(br s, 1H),7.16(s,1H),7.01(br s,1H),6.68(t,J＝55.9Hz,1H),6.28(dd,J＝8.9,2.4 Hz,1H),6.00(d,J＝2.4Hz,1H),4.97-4.86(m,1H),4.60-4.49(m,2H),4.39- 4.28(m,4H),3.94-3.88(m,1H),3.56-3.48(m,1H),3.25-3.16(m,1H),2.22- 2.11(m,1H),2.01-1.86(m,3H)。

example 107(S) -2- ((2- ((S) -4- (difluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propanamide 107

Step 1: (S) -2- ((2- ((S) -4- (difluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid methyl ester

Reacting (S) -2- ((2-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid methyl ester (414mg,1.00mmol), (S) -4-difluoromethyloxazolidin-2-one (143mg,1.05mmol), cuprous iodide (38mg,0.20mmol), trans-N, N' -dimethylcyclohexane-1, 2-diamine (60mg,0.40 mmol)A mixture of l) and potassium carbonate (280mg,2.00mmol) in dioxane (8.0M L) was degassed with argon under sonication and the reaction mixture was heated at 100 ℃ for 6 hours the resulting mixture was diluted with water and extracted with ethyl acetate the combined organic extracts were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo and the crude residue was purified by flash chromatography on silica gel (solvent gradient: 0-60% ethyl acetate/cyclohexane) to give 386mg (91%) of the title compound as a white solid L CMS ESI, [ M + H ]: M + H]⁺＝424。

Step 2: (S) -2- ((2- ((S) -4- (difluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propanamide

Reacting (S) -2- ((2- ((S) -4- (difluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

A solution of-9-yl) oxy) methyl propionate (386mg,0.91mmol) in ammonia solution (10m L, 7N in methanol, 70mmol) was stirred at room temperature for 18 h the reaction mixture was evaporated in vacuo and the resulting white solid triturated with dichloromethane and dried in vacuo to give 325mg (87%) of 107 as a white solid L CMS (ESI)_T(min)＝3.32,[M+H]⁺409, method a;¹H NMR(400MHz,DMSO-d₆)8.22(d,J＝9.0Hz,1H),7.18(s,1H),7.57(br s,1H),7.30-7.25(m,2H),6.88-6.58(m,2H),5.04-4.92(m,1H),4.68-4.53(m,3H),4.45-4.37(m,4H),1.44(d,J＝6.3Hz,3H)。

example 108(S) -1- (2- ((S) -4- (fluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) pyrrolidine-2-carboxamide 108

Step 1: (R) -1- ((tert-butyldimethylsilyl) oxy) -3-fluoropropan-2-ol

Tert-butyldimethylchlorosilane (1.60g,10.63mmol) was added to a solution of (R) -3-fluoropropane-1, 2-diol (1.00g,10.6mmol), triethylamine (1.93m L, 13.8mmol) and catalytic amounts of 4- (dimethylamino) pyridine in dichloromethane, and the reaction mixture was warmed to room temperature and stirred at room temperature for 16 h, the reaction mixture was diluted with water and extracted with dichloromethane, the combined organic fractions were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo, and the resulting crude residue was purified by flash chromatography on silica gel (solvent gradient: 0-40% ethyl acetate/cyclohexane) to give 1.80g (81%) of the title compound as a colorless oil.¹H NMR(400MHz,CDCl₃)4.45-4.36(m,1H),4.34 -4.25(m,1H),3.87-3.73(m,1H),3.66-3.56(m,2H),2.30(d,J＝6.0Hz,1H), 0.82(s,9H),0.00(s,6H)。

Step 2: (S) -2-azido-3-fluoropropoxy) -tert-butyldimethylsilane

Trifluoromethanesulfonic anhydride (1.67m L, 9.93mmol) was added dropwise to a solution of (R) -1- ((tert-butyldimethylsilyl) oxy) -3-fluoropropan-2-ol (1.80g,8.60mmol) and pyridine (1.2m L, 13.8mmol) in dichloromethane and the reaction mixture was stirred at-20 ℃ for 20 minutes, then at 0 ℃ for 30 minutes, the reaction mixture was diluted with 0.5N aqueous HCl and extracted with dichloromethane, the combined organic extracts were dried over magnesium sulfate, filtered and evaporated in vacuo, the residue was dissolved in N, N-dimethylformamide (5.0m L) and sodium azide (1.68g,25.9mmol) was added, the reaction mixture was stirred at room temperature for 2 hours, the resulting mixture was diluted with water and extracted with ethyl acetate, the combined organic extracts were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo to give the crude title compoundThe compound, which can be used without further purification.¹H NMR(400MHz,CDCl₃)4.58-4.26(m,2H),3.75-3.63(m, 2H),3.62-3.46(m,1H),0.80(s,9H),0.00(s,6H)。

And step 3: (S) -1- ((tert-butyldimethylsilyl) oxy) -3-fluoropropan-2-amine

Palladium hydroxide (400mg, 20% on carbon) was added to a solution of ((S) -2-azido-3-fluoropropoxy) -tert-butyldimethylsilane (crude, assumed to be 8.60mmol) in ethyl acetate (15m L) and methanol (5.0m L) and the reaction mixture was stirred under a hydrogen balloon for 16 hours the resulting mixture was filtered, fresh palladium hydroxide (400mg, 20% on carbon) was added and the reaction was stirred under a hydrogen balloon for another 16 hours the resulting mixture was filtered and the filtrate evaporated in vacuo to give the title compound as the product, an approximately 2:1 mixture of the starting materials which was used on without purification.

And 4, step 4: (S) -4-fluoromethyl oxazolidin-2-one

HCl/dioxane (4N,2.0m L, 8.00mmol) was added to a solution of (S) -1- ((tert-butyldimethylsilyl) oxy) -3-fluoropropan-2-amine (crude, assumed to be 8.60mmol) in methanol (3.0m L) and the resulting mixture was stirred at room temperature for 2 hours the reaction mixture was evaporated under vacuum, the resulting residue was dissolved at 0 ℃ in a mixture of toluene (20m L) and KOH (2.89g,51.6mmol, 12.5% aqueous solution), phosgene (13.6m L, 20% in toluene) was added dropwise to the mixture, the cooling bath was removed and the resulting mixture was stirred for 1 hour, the reaction mixture was evaporated under vacuum, the resulting residue was extracted with hot technical methylated spirit, the filtrate was evaporated under vacuum, the resulting residue was purified by silica gel flash chromatography (solvent gradient: 50-100% ethyl acetate/cyclohexane) to give 450mg (44%, 3 steps) of the title compound as an off-white solid.¹H NMR(400MHz,CDCl₃)5.69(br s,1H),4.59-4.42(m,2H), 4.42-4.32(m,1H),4.25-4.08(m,2H)。

And 5: (S) -3- (9-bromo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4- (fluoromethyl) oxazolidin-2-one and (S) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4- (fluoromethyl) oxazolidin-2-one

Reacting 9-bromo-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

(722mg,1.85mmol), (S) -4-fluoromethyloxazolidin-2-one (220mg,1.85mmol), 3,4,7, 8-tetramethyl-1, 10-phenanthroline (131mg,0.55mmol), Cu (OAc)₂.H₂A mixture of O (74mg,0.37mmol), potassium carbonate (510 mg,3.70mmol) and dioxane (6.0ml) was sealed in a tube and the mixture was degassed with argon under sonication conditions the reaction mixture was heated at 100 ℃ for 72 hours the resulting reaction mixture was diluted with 15% aqueous ammonia and extracted with ethyl acetate the combined organic extracts were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo the crude residue was purified by silica gel flash chromatography (solvent gradient: 0-100% ethyl acetate/cyclohexane) to give 390mg (53%) of the title compound (an about 2:1 mixture of 9-Br and 9-I product): L CMS (ESI): M + H]⁺＝382/384/430；¹H NMR(400MHz,CDCl₃) 8.22(d,J＝9.3Hz,0.7H),8.05(d,J＝8.8Hz,0.3H),7.43-7.37(m,0.6H), 7.29(s,1.2H),7.23-7.18(m,1.2H),5.03-4.66(m,3H),4.60(t,J＝8.5Hz,1H), 4.54(dd,J＝8.6,4.3Hz,1H),4.47-4.43(m,2H),4.37-4.33(m,2H)。

Step 6: (S) -1- (2- ((S) -4- (fluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxygen and nitrogenHetero compound

-9-yl) pyrrolidine-2-carboxamide

Reacting (S) -3- (9-bromo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4- (fluoromethyl) oxazolidin-2-one and (S) -3- (9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-2-yl) -4- (fluoromethyl) oxazolidin-2-one (270mg,2:1 mixture, about 0.71mmol), L-proline (163 mg,1.41mmol), cuprous iodide (27mg,0.14mmol) and tripotassium phosphate (300mg,1.41mmol) in dimethyl sulfoxide (2.0m L) were degassed with argon under ultrasonic conditions the reaction mixture was heated at 100 ℃ for 2 hours the resulting mixture was cooled to room temperature and diluted with dimethyl sulfoxide (1.0m L) ammonium chloride (227mg,4.20mmol), triethylamine (0.98m L, 7.10mmol), 1- [ bis (dimethylamino) methylene ] chloride]-1H-1,2, 3-triazolo [4,5-b]Pyridinium 3-oxide hexafluorophosphate (1.61g, 4.20mmol) and the reaction mixture stirred at room temperature for 30 minutes the resulting mixture was diluted with water and extracted with ethyl acetate the combined organic extracts were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo and the crude residue was purified by flash chromatography on silica gel (solvent gradient: 0-10% methanol/DCM) to give 150mg (51%) of 108. L CMS (ESI): M + H]⁺＝416；¹H NMR(400MHz,d₆-DMSO)8.02 d,J＝8.7Hz,1H),7.36(br s,1H),7.16(s,1H),7.01(br s,1H),6.28(dd,J＝7.7, 2.5Hz,1H),6.00(d,J＝2.5Hz,1H),4.97(ddd,J＝48.5,10.0,2.8Hz,1H),4.78 -4.53(m,3H),4.40-4.27(m,5H),3.94-3.88(m,1H),3.56-3.48(m,1H),3.25 -3.16(m,1H),2.22-2.11(m,1H),1.99-1.87(m,3H)。

Example 109: (S) -2- ((2- ((S) -4- (fluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propanamide 109

Step 1: (S) -2- ((2- ((S) -4- (fluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionic acid methyl ester

Reacting (S) -2- ((2-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) methyl propionate (414mg,1.00mmol), (S) -4-fluoromethylooxazolidin-2-one (123mg,1.05mmol), cuprous iodide (38mg,0.20mmol), trans-N, N' -dimethylcyclohexane-1, 2-diamine (60mg,0.40 mmol) and potassium carbonate (280mg,2.00mmol) in dioxane (8.0M L) a mixture was degassed with argon under ultrasonic conditions and the reaction mixture was added at 100 ℃ for 18 hours the resulting mixture was diluted with water and extracted with ethyl acetate the combined organic extracts were washed with brine, dried over magnesium sulfate, filtered and evaporated in vacuo the crude residue was purified by silica gel flash chromatography (solvent gradient: 0-80% ethyl acetate/cyclohexane) to give 305mg (75%) of the title compound as a white solid L (esi): L cms [ M + H: [ esi ], [ M + H ])]⁺＝ 406。

Step 2: (S) -2- ((2- ((S) -4- (fluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propanamide

Reacting (S) -2- ((2- ((S) -4- (fluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

A solution of-9-yl) oxy) methyl propionate (300mg,0.74mmol) in ammonia solution (10m L, 7N in methanol, 70.0mmol) was stirred at room temperature for 18 hours the resulting white solid was filtered and evaporated in vacuo to give 190mg (66%) of 109 as a white solid L CMS (ESI): R_T(min)＝2.91,[M+H]⁺391, method a;¹H NMR(400MHz,DMSO-d₆) 8.17(d,J＝8.9Hz,1H),7.54(br s,1H),7.27(s,1H),7.25(br s,1H),6.71(dd,J ＝9.0,2.5Hz,1H),6.50(d,J＝2.5Hz,1H),5.00(ddd,J＝48.4,9.8,2.6Hz,1H),4.83-4.57(m,4H),4.45-4.37(m,5H),1.43(d,J＝6.6Hz,3H)。

example 901PI3K binding assay

The PI3K binding assay is intended to determine the biochemical potency of small molecule PI3K inhibitors. The PI3K lipid kinase reaction was performed in the presence of PIP2:3PS lipid substrate (Promega # V1792) and ATP. After termination of the kinase reaction, Promega ADP-Glo was used^TM(Promega # V1792) assay detects whether the kinase reaction converts ATP to ADP by phosphorylation of phospholipid substrates. As shown in table 5, the reaction was performed using the following conditions for each PI3K isoform.

Table 5.

After 120 minutes of reaction, the kinase reaction was terminated. Any ATP remaining after the reaction is depleted leaving only ADP. Kinase detection reagents are then added to convert ADP to ATP, which is used in the coupled luciferin/luciferase reaction. The luminescence output was measured and correlated with kinase activity.

All reactions were performed at room temperature for each PI3K isoform, 3 μ l of a mixture of enzyme/lipid substrate solutions (1:1) was added to 384-well white assay plates containing only 50n L test compound or DMSO (Perkin Elmer #6007299) only for untreated controls by adding 2 μ L ATP/MgCl₂The reaction was started. The kinase reaction buffer contained 50mM HEPES, 50mM NaCl, 3mM MgCl₂0.01% BSA, 1% DMSO, andthe reaction was stopped by the addition of 10 μ L ADP-Glo reagent.plates were read in a Perkin Elmer Envision system using a luminescence pattern.A 10-point dose response curve was generated for each test compound.Ki values for each compound were determined using the Morrison equation.

Binding assays initial polarization experiments were performed on analysis HT 96-384(Molecular Devices Corp, Sunnyvale, Calif.) samples for fluorescence polarization affinity measurements were prepared by serial 1:3 dilutions of p110 α PI3K (Upstate Cell Signaling Solutions, Charlottesville, Va.) starting in polarization buffer (10mM Tris pH 7.5, 50mM NaCl, 4mM MgCl. sub.₂0.05% Chaps and 1mM DTT) to a final concentration of 20. mu.g/m L) to a final concentration of 10mM PIP₂(Echelon-inc., Salt L ake City, UT.) after incubation for 30 minutes at room temperature, the reaction was stopped by the addition of GRP-1 and PIP3-TAMRA probes (Echelon-inc., Salt L ake City, UT.) at final concentrations of 100nM and 5nM, respectively

Rhodamine fluorophores were read (lambda) with standard cut-off filters in (PerkinElmer, Wellesley, MA.)_Excitation＝530nm；λ_Launching590 nm). The fluorescence polarization values were plotted as a function of protein concentration. EC (EC)₅₀The values are obtained as follows: use of

Software (Synergy software, Reading, PA) fits data to a four parameter equation. This experiment also determined the protein concentration suitable for subsequent inhibitor competition experiments.

Inhibitor IC₅₀The value was determined by mixing 0.04mg/m L p110 α PI3K (final concentration) with PIP₂(10mM final concentration) were added together to wells containing 1:3 serial dilutions of the antagonist in ATP (Cell Signaling technology, Inc., Danvers, MA)/polarizing buffer at a final concentration of 25 mM after 30 minutes incubation at room temperature, the reaction was stopped by the addition of GRP-1 and PIP3-TAMRA probes (Echelon-Inc., Salt L ake City, UT.) at final concentrations of 100nM and 5nM, respectively. Black low capacity in 384 wells

Rhodamine fluorophores were read (lambda) with standard cut-off filters in (PerkinElmer, Wellesley, MA.)_Excitation＝530nm；λ_Launching590 nm). Plotting fluorescence polarization values as a function of antagonist concentration and IC₅₀The values were obtained by fitting the data to a four parameter equation in the Assay Explorer software (MD L, San Ramon, CA.).

Alternatively, inhibition of PI3K was determined in a radioactive assay using purified recombinase and ATP at a concentration of 1 μ M (micromolar). Compounds were serially diluted in 100% DMSO. The kinase reaction mixture was incubated at room temperature for 1h and the reaction was stopped by adding PBS. IC was then determined using sigmoidal dose-response curve fitting (variable slope)₅₀The value is obtained.

Co-crystallography of example 902 with p110 α (alpha)

N-terminally truncated p110 α (alpha) was generated according to Chen et al and Nacht et al (Chen, P., Y. L. Deng, S.Bergqvist, M.D.Falk, W. L iu, S.Timofevski and A.Brooun "Engineering of isolated p110 alpha. supplement of PI3Kalpha permit crystallization and providesa platform for structural-based drug device," (2014) Protein Sci 23(10): 1332-.

In the presence of the subject compounds, standard protocols were used to produce crystals. The harvested crystals were stored by immersion in liquid nitrogen for diffraction data collection and mounted on a synchrotron beam line producing monochromatic X-rays. Diffraction data was collected, reduced and combined using standard protocols. The crystal unit cells and space groups are isomorphous to those previously reported (Nacht, 2013; Chen, 2014). Place the project compounds in electron density maps and perform crystallographic refinements to 2.36-

Resolution limit of (2).

Example 903Selective inhibition of mutant PI3K α (alpha)

The ability of the compounds of the invention to act preferentially on cells containing mutant PI3K α (alpha) was determined by measuring inhibition of the PI3K pathway in SW48 isogenic cell lines PI3K α wild type (parental), helical domain mutant E545K and kinase domain mutant H1047R the following assay is intended to determine the cellular potency and mutant selectivity of small molecule PI3K α inhibitors using isogenic cell lines expressing PI3K α WT, PI3K α mutant E545K/+ (horizonn Discovery 103-001) or PI3K α mutant H1047R/+ (horizonn Discovery 103-005) measuring pPRAS40 inhibited by PI3K α in each cell line 24 hours after compound treatment the mutation selectivity of PI3 38964 inhibitor was determined by WT 3K α versus E545K and WT 545 versus EC 10463 in H10463-1047R cell lines₅₀And (5) determining the efficacy ratio.

Cell culture cell lines were maintained in a cell culture chamber at 37 ℃ and 5% in cell culture medium containing RPMI1640 (prepared in Genentech), 10% FBS (Gibco 16140-.

Measurement operation: cells were harvested and plated in 384-well tissue culture treated assay plates (Greiner cat #781091) and at 37 ℃ with 5% CO₂Incubate overnight. Three cell lines (WT, E545K and H1047R) were plated in parallel and assayed. The next day, test compounds were serially diluted in dimethyl sulfoxide (DMSO) and added to the cells (final DMSO concentration of 0.5%). The cells were then incubated at 37 ℃ and 5% CO₂After 24 hours of incubation, the cells were lysed and the level of pPRAS40 was measured using the Meso-Scale custom pPRAS 40384 w assay kit (Meso-Scale Discovery, cat # L21 CA-1). cell lysate was added to assay plates pre-coated with antibodies against phosphorylated PRAS 40. phosphorylated PRAS40 in the sample was allowed to bind to the capture antibody overnight at 4 ℃. detection antibody (anti-total PRAS40 labeled with electrochemiluminescent SU L FO-TAG) was added to the bound lysateAnd incubated at room temperature for 1 hour. The MSD read buffer was added so that when a voltage was applied to the plate electrode, the label bound to the electrode surface emitted light. The MSD fan instrument measures the intensity of the light and quantitatively measures the amount of phospho-PRAS 40 in the sample. The percent inhibition of phosphorylation of PRAS40 by different concentrations of test compound was calculated relative to untreated controls. EC calculation Using a 4-parameter logistic non-Linear regression dose-response model₅₀The value is obtained.

Statistical analysis: EC (EC)₅₀Values represent geometric means of a minimum of 4 independent experiments. All statistics were performed using KaleidaGraph software (version 4.1.3). Student's t-test was performed using unpaired data with the same variance to compare activity against mutant and wild type cells. P<0.05 was considered significant.

Example 904In vitro cell viability assay

Cells (1500 per well) were seeded in 384-well plates for 16 hours. On day 2, 9 serial 1:3 compound dilutions in DMSO were prepared in 96-well plates. The compounds were then further diluted into growth medium using a Rapidplate robot (Zymark Corp.). The diluted compounds were then added to quadruplicate wells of 384-well cell plates and incubated at 37 ℃ and 5% CO₂And (4) incubation. After 4 days, the relative number of viable cells was measured by luminescence using Cell Titer-glo (promega) according to the manufacturer's instructions and read on a Wallac Multilabel Reader (Perkin-Elmer). EC was calculated using prism6.0 software (GraphPad)₅₀The value is obtained.

Example 905In vivo mouse tumor xenograft efficacy

Mice: female severe combined immunodeficiency mice (

C.B-17/Charles River Holister) may be at 10 to 11 weeks of age, with body weights ranging from 15.8 to 25.09 grams on study day 0 animals were fed water ad libitum (reverse osmosis, 1ppm Cl) and NIH 31modified and Irradiated laboratory diet consisting of 18.0% crude protein, 5.0% crude fat and 5.0% crude fiber (NIH 31modified and Irradated L ab)

) In irradiated A L PHA-

bed-

The animal care and use procedures of Genentech are approved by the international society for laboratory animal husbandry evaluation certification (AAA L AC), which ensures compliance with accepted laboratory animal care and use standards.

Tumor implantation starting with cancer cells xenograft was initiated by culturing KP L4 breast cancer cells in RPMI1640 medium supplemented with 10% fetal bovine serum, 2mM glutamine, 100 units/m L penicillin, 100. mu.g/m L streptomycin sulfate, and 25. mu.g/m L gentamicin, harvesting the cells during exponential phase growth and growing at 3 × 10⁶The concentration of mice was resuspended in matrigel HBSS. Tumor cells were implanted subcutaneously into the right breast fat pad when the average size was approximately 150 to 200mm³Tumor growth was monitored at target range. 14 days after tumor implantation, designated as study day 0, mice were divided into 10 groups of 10 mice each, where for 13-3399C (Compound 102) the individual tumor volumes were 128-235mm³For 13-3399D (Compound 103), the individual tumor volumes were 130-³. The group mean tumor volume for study 13-3399C was 171-180mm³Group mean tumor volume of 200- & ltwbr & gt 214mm for studies 13-3399D³. The volume was calculated using the formula:

tumor volume (mm)³)＝(w²x l)/2, wherein w is the tumor width in mm and l is the tumor length in mm. Assuming that 1mg equals 1mm³The tumor weight can be estimated.

Therapeutic agent pharmaceutical doses were prepared in 0.5% methylcellulose: 0.2% tween 80 in deionized water ("vehicle") and stored at 4 ℃, compound 102 was formulated in a nanosuspension in MCT and stored at 4 ℃, compound 103 was prepared in MCT weekly and stored at 4 ℃, all doses were formulated to deliver the mg/kg dose at a volume of 0.1m L per 22 grams body weight (4m L/kg).

Treatment: all doses were scaled according to the body weight of the individual animals and provided by the routes indicated.

End point: tumor volumes were measured in two dimensions (length and width) using an Ultra Cal IV caliper (Model 5410111; Fred v. fowler company) as follows: tumor volume (mm)³) Length × width²) ×.5 and analyzed using Excel version 11.2 (Microsoft Corporation.) linear mixing (L ME) modeling method was used to analyze repeated measurements of tumor volume from the same animals over time (Pinheiro, j. et al (2009); Tan, n.et al (2011) clin.cancer res.17(6): 1394-1404.) this method compromised both repeated measurements and appropriate withdrawal due to death of any non-treatment related animals before the end of the study_Medicine/AUC_Media). Using this formula, a value of 100% TGI indicates tumor arrest,>1% but<A value of 100% TGI indicates a delay in tumor growth, and>TGI values of 100% indicate tumor regression. Partial Response (PR) in animals was defined as tumor regression>50% but<100% of the initial tumor volume. Complete Response (CR) was defined as 100% tumor regression (i.e. no measurable tumor) on any day during the study.

Toxicity: animals were weighed twice a week and the Adventurer was used for body weight during the study

AV812 scale (Ohaus Corporation). The percent weight change was calculated as follows: change in body weight (%) - (weight)_{New day}-weight_{Day 0}) Weight/weight_{Day 0}]× 100, mice are often observed for any significant signs of adverse treatment-related side effects and clinical signs of toxicity are recorded when observed acceptable toxicity is defined as a group average Body Weight (BW) loss of less than 20% and no more than 1 treatment-related (TR) death out of 10 treated animals over the course of the study.

Although the foregoing invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, the description and example should not be construed as limiting the scope of the invention. The entire disclosures of all patent and scientific literature cited in this application are expressly incorporated by reference into this application.

Claims (13)

1. A compound selected from:

(S) -2- ((2- ((S) -2-oxo-4- (trifluoromethyl) oxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionamide;

(S) -1- (2- ((S) -4- (difluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) pyrrolidine-2-carboxamide;

(S) -2- ((2- ((S) -4- (difluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionamide;

(S) -1- (2- ((S) -4- (fluoromethyl)) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) pyrrolidine-2-carboxamide; and

(S) -2- ((2- ((S) -4- (fluoromethyl) -2-oxooxazolidin-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d ] s][1,4]Oxazazem

-9-yl) oxy) propionamide.

2. A pharmaceutical composition consisting of a compound of claim 1 and a pharmaceutically acceptable carrier, glidant, diluent, or excipient.

3. The pharmaceutical composition of claim 2, wherein the pharmaceutically acceptable carrier, glidant, diluent, or excipient is selected from the group consisting of silicon dioxide, powdered cellulose, microcrystalline cellulose, metal stearates, sodium aluminosilicate, sodium benzoate, calcium carbonate, calcium silicate, corn starch, magnesium carbonate, asbestos-free talc, stearowet C, starch 1500, magnesium lauryl sulfate, magnesium oxide, and combinations thereof.

4. A process for preparing a pharmaceutical composition comprising combining a compound of claim 1 with a pharmaceutically acceptable carrier, glidant, diluent, or excipient.

5. Use of a compound of claim 1 in the manufacture of a medicament for treating cancer in a patient, wherein the cancer is selected from breast cancer and non-small cell lung cancer.

6. The use of claim 5, wherein the cancer is breast cancer.

7. The use of claim 6, wherein the breast cancer is estrogen receptor positive breast cancer.

8. The use of claim 6, wherein the breast cancer subtype is basal or tubular epithelial.

9. The use of claim 6, wherein the cancer expresses a PIK3CA mutant selected from the group consisting of E542K, E545K, Q546R, H1047L, and H1047R.

10. The use of claim 6 wherein the cancer expresses a PTEN mutant.

11. The use of claim 6, wherein the cancer is HER2 positive.

12. The use of claim 6, wherein the patient is HER2 negative, estrogen receptor negative, and progesterone receptor negative.

13.A kit for therapeutic treatment of breast cancer, comprising:

a) the pharmaceutical composition of claim 2; and

b) instructions for use in the therapeutic treatment of breast cancer.

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